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Xing Zhu,1,* Hao Wang,2,* Longbin Zheng,1 Zhaoyu Zhong,1 Xuesong Li,1 Jing Zhao,3 Jiayuan Kou,1 Yueqing Jiang,1 Xiufeng Zheng,1 Zhongni Liu,1 Hongxia Li,1 Wenwu Cao,4,5 Ye Tian,1,6 You Wang,2 Liming Yang1 1Department of Pathophysiology, Harbin Medical University, Harbin, People’s Republic of China; 2Materials Physics and Chemistry Department, Harbin Institute of Technology, Harbin, People’s Republic of China; 3Blood Transfusion Department, Jining No 1 People’s Hospital, Jining, People’s Republic of China; 4Laboratory of Sono- and Photo-theranostic Technologies, Harbin Institute of Technology, Harbin, People’s Republic of China; 5Materials Research Institute, The Pennsylvania State University, University Park, PA, USA; 6Division of Cardiology, The First Affiliated Hospital, Harbin Medical University, Harbin, People’s Republic of China *These authors contributed equally to this work Abstract: Atherosclerosis (AS) is the most vital cardiovascular disease, which poses a great threat to human health. Macrophages play an important role in the progression of AS. Photodynamic therapy (PDT) has emerged as a useful therapeutic modality not only in the treatment of cancer but also in the treatment of AS. The purpose of this study was to determine the molecular mechanisms underlying the activity of PDT, using mesoporous-silica-coated upconversion fluorescent nanoparticles encapsulating chlorin e6 (UCNPs-Ce6) in the induction of apoptosis in THP-1 macrophages. Here, we investigated the ability of UCNPs-Ce6-mediated PDT to induce THP-1 macrophage apoptosis by facilitating the induction of reactive oxygen species (ROS) and regulation of mitochondrial permeability transition pore (MPTP) to depolarize mitochondrial membrane potential (MMP). Both Bax translocation and the release of cytochrome C were examined using immunofluorescence and Western blotting. Our results indicated that the levels of ROS were significantly increased in the PDT group, resulting in both MPTP opening and MMP depolarization, which led to apoptosis. In addition, immunofluorescence and Western blotting revealed that PDT induced both Bax translocation and the release of cytochrome C, as well as upregulation of cleaved caspase-9, cleaved caspase-3, and cleaved poly(ADP-ribose) polymerase. Therefore, we demonstrated that UCNPs-Ce6-mediated PDT induces apoptosis in THP-1 macrophages via ROS bursts. The proapoptotic factor Bax subsequently translocates from the cytosol to the mitochondria, resulting in the MPTP opening and cytochrome C release. This study demonstrated the great potential of UCNPs-Ce6-mediated PDT in the treatment of AS. Keywords: atherosclerosis, photodynamic therapy, fluorescent nanoparticles, reactive oxygen species, apoptosis, macrophages

Xuesong Li,1,* Lei Gao,2,* Longbin Zheng,1 Jiayuan Kou,1 Xing Zhu,1 Yueqing Jiang,1 Zhaoyu Zhong,1 Juhua Dan,1 Haobo Xu,3 Yang Yang,3 Hong Li,1 Sa Shi,1 Wenwu Cao,4,5 Yajun Zhao,1 Ye Tian,1,3 Liming Yang1 1Department of Pathophysiology, Harbin Medical University, Harbin, People’s Republic of China; 2Electron Microscopy Centre, Harbin Medical University, Harbin, People’s Republic of China; 3Division of Cardiology, The First Affiliated Hospital, Harbin Medical University, Harbin, People’s Republic of China; 4Laboratory of Sono- and Photo-theranostic Technologies, Harbin Institute of Technology, Harbin, People’s Republic of China; 5Materials Research Institute, The Pennsylvania State University, University Park, PA, USA *These authors contributed equally to this work Purpose: To investigate the sonoactivity of hypericin (HY), together with its sonodynamic effect on THP-1 macrophages and the underlying mechanism.Materials and methods: CCK-8 was used to examine cell viability. Confocal laser scanning microscopy was performed to assess the localization of HY in cells, reactive oxygen species (ROS) generation, and opening of the mitochondrial permeability transition pore (mPTP) after different treatments. Apoptosis was analyzed using Hoechst–propidium iodide and transmission electron microscopy. Mitochondrial membrane potential (ΔΨm) collapse was detected via fluorescence microscopy. Lipoprotein oxidation was determined in malondialdehyde (MDA) assays. Western blotting was conducted to determine the translocation of BAX and cytochrome C and the expression of apoptosis-related proteins.Results: HY was sublocalized among the nuclei and the mitochondria, endoplasmic reticulum, Golgi apparatus, and lysosome in the cytosol of THP-1 macrophages. Under low-intensity ultrasound irradiation, HY significantly decreased cell viability and induced apoptosis. Furthermore, greater ROS generation, higher MDA levels, and greater ΔΨm loss were observed in the sonodynamic therapy (SDT) group. Both ROS generation and MDA levels were significantly reduced by the ROS scavenger N-acetyl cysteine (NAC) and the singlet oxygen scavenger sodium azide. Most of the loss of ΔΨm was inhibited by pretreatment with NAC, sodium azide, and the mPTP inhibitor cyclosporin A (CsA). mPTP opening was induced upon SDT but was reduced by pretreatment with bongkrekic acid, 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid disodium, CsA, and NAC. Western blot analyses revealed translocation of BAX and cytochrome C, downregulated expression of Bcl-2, and upregulated expression of cleaved caspase-9, cleaved caspase-3, and cleaved poly(ADP-ribose) polymerase in the SDT group, which were reversed by NAC.Conclusion: HY mediated SDT-induced apoptosis in THP-1 macrophages via ROS generation. Then, the proapoptotic factor BAX translocated from the cytosol to the mitochondria, increasing the ratio of BAX/Bcl-2, and the mPTP opened to release cytochrome C. This study demonstrated the great potential of HY-mediated SDT for treating atherosclerosis. Keywords: apoptosis, hypericin, sonodynamic therapy, mitochondria–caspase pathway, atherosclerosis

Objective: To study the influence of distal tibiofibular synostosis on ankle function.Methods: From October 1998 to October 2004, a total of 281 consecutive patients underwent operations because of ankle fractures or distal fractures of the tibia and fibula. Distal tibiofibular synostosis occurred after operation in 8 patients. The duration of follow-up averaged 20.6 months (14-44 months). The ankle function was assessed on the basis of functional rating system described by Mazur.(1)Results: According to Mazur's ankle evaluation system, 4 patients achieved an excellent result, 2 a good result and 2 a fair result. The dorsiflexion of the synostosis ankle reduced by 8.26 degrees as compared with that of the contralateral ankle, and there was little influence on the plantar flexion. All the patients had a normal gait.Conclusion: The distal tibiofibular synostosis after the operation of ankle fractures or distal fractures of the tibia and fibula usually gives rise to few symptoms and needs no specific treatment. [ABSTRACT FROM AUTHOR]

FMRFamide, a member of the neuropeptide family, is involved in numerous physiological processes. FMRFamide-activated sodium channels (FaNaCs) are a family of non-voltage-gated, amiloride-sensitive, Na + -selective channels triggered by the neuropeptide FMRFamide. In the present study, the full-length cDNA of the FaNaC receptor of Sepiella japonica (SjFaNaC) was cloned. The cDNA of SjFaNaC was 3004 bp long with an open reading frame (ORF) of 1812 bp, encoding 603 amino acid residues with no signal peptide at the N-terminus. Sequence analysis indicated that SjFaNaC shared a high identity with other cephalopods FaNaCs and formed a sister clade with bivalves. The protein structure was predicted using SWISS-MODEL with AcFaNaC as the template. Quantitative real-time PCR (qRT-PCR) revealed that SjFaNaC transcripts were highly expressed in both female and male reproductive organs, as well as in the optic lobe and brain of the central nervous system (CNS). Results of in situ hybridisation (ISH) showed that SjFaNaC mRNA was mainly distributed in the medulla and deep retina of the optic lobe and in both the supraesophageal and subesophageal masses of the brain. Subcellular localisation indicated that the SjFaNaC protein was localised intracellularly and on the cell surface of HEK293T cells. In summary, these findings may lay the foundation for future exploration of the functions of SjFaNaC in cephalopods., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Large yellow croaker (Larimichthys crocea) is the most productive marine fish in China. Cryptocaryon irritans is an extremely destructive parasite that causes great economic losses in large yellow croaker aquaculture industry. Therefore, it is very necessary to study the immune response of large yellow croaker in response to C. irritans infection. In this study, the transcriptomic profiles of large yellow croaker were sequenced and analyzed in the brain and head kidney at 72 h after C. irritans infection. Cytokines and chemokines related terms were significantly enriched based on the GO enrichment of down-regulated differentially expressed genes (DEGs) from the head kidney. Meanwhile, cytokine-cytokine receptor interaction was significantly enriched based on the KEGG enrichment of up-regulated DEGs from the brain and down-regulated DEGs from the head kidney, respectively. Moreover, the majority of inflammation-related DEGs were significantly up-regulated in the brain, but distinctly down-regulated in the head kidney. These results showed that the brain and head kidney might play different roles against C. irritans infection, and the inflammatory response of large yellow croaker may be restrained during C. irritans infection. Taken together, the transcriptomic analyses will be helpful to more comprehensively understand the immune mechanism of teleost against C. irritans infection, and provide a theoretical basis for the prevention and treatment of Cryptosporidiosis., (Copyright © 2023. Published by Elsevier Ltd.)

Objective: To examine the radiomics model based on high-resolution T2WI and diffusion weighted imaging (DWI) in predicting microsatellite stability in patients with stage Ⅱ and Ⅲ rectal cancer. Methods: From February 2016 to October 2020, 175 patients with stage Ⅱ and Ⅲ rectal cancer who met the inclusion criteria were retrospectively collected. There were 119 males and 56 females, aged (63.9±9.4) years (range: 37 to 85 years), including 152 patients with microsatellite stability and 23 patients with microsatellite instability. All patients were randomly divided into the training group ( n =123) and the validation group ( n =52) with a ratio of 7∶3. The region of interest was labeled on the T2WI and DWI images of each patient using the ITK-SNAP software, and PyRadiomics was used to extract seven kinds of radiomics features. After removing redundant features and normalizing features, the least absolute shrinkage and selection operation were used for feature selection. One clinical model, three radiomics models and one clinical-radiomics model were constructed in the training group based on a support vector machine. The area under receiver operating characteristic curve (AUC), sensitivity, specificity, and accuracy were used to evaluate the performance of the models in the verification group. Results: Three clinical features (age, degree of tumor differentiation, and distance from the lower edge of the tumor to the anal edge) and six radiomics features (two DWI-related features and four T2WI-related features) most related to microsatellite status of rectal cancer patients were selected. The AUC of the clinical-radiomics model in the training group was 0.95. In the validation group, the AUC was 0.81, better than the clinical model (0.68, Z =0.71, P =0.04), and equivalent to the T2WI+DWI model (0.82, Z =0.21, P =0.83). Conclusions: Radiomic features based on preoperative T2WI and DWI were related to microsatellite stability in patients with stage Ⅱ and Ⅲ rectal cancer and showed a high classification efficiency. The model based on the features provided a noninvasive and convenient tool for preoperative determination of microsatellite stability in rectal cancer patients.

Stronger intrinsic Warburg effect and resistance to chemotherapy are the responses to high mortality of renal cell carcinoma (RCC). Pyruvate kinase M2 (PKM2) plays an important role in this process. Promoting PKM2 conversion from dimer to tetramer is a critical strategy to inhibit Warburg effect and reverse chemotherapy resistance. Herein, a PKM2 allosteric converter (PAC) is constructed based on the "in vivo self-assembly" strategy, which is able to continuously stimulate PKM2 tetramerization. The PAC contains three motifs, a serine site that is protected by enzyme cleavable β-N-acetylglucosamine, a self-assembly peptide and a AIE motif. Once PAC nanoparticles reach tumor site via the EPR effect, the protective and hydrophilic β-N-acetylglucosamine will be removed by over-expressed O-GlcNAcase (OGA), causing self-assembled peptides to transform into nanofibers with large serine (PKM2 tetramer activator) exposure and long-term retention, which promotes PKM2 tetramerization continuously. Our results show that PAC-induced PKM2 tetramerization inhibits aberrant metabolism mediated by Warburg effect in cytoplasm. In this way, tumor proliferation and metastasis behavior could be effectively inhibited. Meanwhile, PAC induced PKM2 tetramerization impedes the nuclear translocation of PKM2 dimer, which restores the sensitivity of cancer cells to first-line anticancer drugs. Collectively, the innovative PAC effectively promotes PKM2 conversion from dimer to tetramer, and it might provide a novel approach for suppressing RCC and enhancing chemotherapy sensitivity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

As one of the key components of innate immune system, piscidins are likely to play pivotal role in the first defense line in fish. Piscidins own multiple resistance activity. A novel piscidin 5-like type 4 was excavated from Larimichthys crocea (termed Lc-P5L4) liver transcriptome immuned by Cryptocaryon irritans, and upregulated at 7 days post infection when secondary bacterial infection occurred. In the study, we characterized the antibacterial activity of Lc-P5L4. The liquid growth inhibition assay detected the recombinant Lc-P5L4 (rLc-P5L) had potent antibacterial activity to Photobacterium damselae. Scanning electron microscope (SEM) observed the cell surface of P. damselae collapsed to form pit, and membrane of some bacteria ruptured after co-incubation with rLc-P5L. Further, transmission electron microscope (TEM) was also employed to observe the intracellular microstructural damage, rLc-P5L4 caused cytoplasm contraction, pores formation and contents leakage. After knowing about its antibacterial effects, the preliminary antibacterial mechanism was also explored, western blot analysis showed rLc-P5L4 could bind to P. damselae through targeting to LPS. Agarose gel eletrophoresis analysis further showed rLc-P5L4 could also penetrate into cells and brought about genome DNA degradation. Therefore, rLc-P5L4 was of potential being a candidate to explore new antimicrobial drug or additive agent, especially to P. damselae., Competing Interests: Declaration of competing interest The paper is co-authored by Li-bing Zheng, Yue-xin Li, Yongquan Su. All the authors agreed to submit the research paper titled “Antibacterial activity study of a novel piscidin 5-like type 4 from Larimichthys crocea” for publication in *Fish and Shellfish Immunology*. This manuscript has not been published or presented elsewhere in part or in entirety and is not under consideration by another journal. We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled., (Copyright © 2023. Published by Elsevier Ltd.)

Objective: To investigate the regulatory effect and mechanism of vitamin D on the local renin-angiotensin system at maternal-fetal interface in the pathological process of preeclampsia (PE)., Methods: The mRNA and protein expression of renin in decidua of normal pregnancy and PE placentas was determined by RT-PCR and Western blot. Normal decidual tissues were treated with active and inactive vitamin D for 48 h in vitro and the expressions of renin and vitamin D deactivating enzyme CYP24A1 were determined by RT-PCR and Western blot. Normal decidual stromal cells and glandular epithelial cells were isolated and purified, and identified by immunocytochemical staining. RT-PCR was used to examine the mRNA of vdr , cyp 27 b 1, cyp 24 a 1, and renin in the two types of cells and in decidual tissue, and the mRNA products were subjected to gel electrophoresis. These two cell types were treated with active and inactive vitamin D in vitro and the expressions of renin and vitamin D deactivating enzyme CYP24A1 were determined by RT-PCR and Western blot. Decidual gland epithelial cells were treated with protein kinase A (PKA) activator forskolin or inhibitor H89 to explore the interaction between PKA pathway and vitamin D in the regulation of renin expression., Results: The expression of renin in PE decidua was significantly higher than that of normal control at transcriptional and translational levels ( P <0.05). Vitamin D treatment could significantly down-regulate the expression of renin in normal decidua tissues ( P <0.05), while it significantly up-regulated CYP24A1 expression ( P <0.001). Decidual stromal cells and gland epithelial cells were successfully isolated from decidual tissue. Compared with that in decidual stromal cells, the mRNA level of vitamin D-related molecules in gland epithelial cells was more similar to that in decidual tissue. Active or inactive vitamin D treatment significantly inhibited the expression of renin in glandular epithelial cells ( P <0.05), but the expression of renin in decidual stromal cells was not affected. However, the treatment of active or inactive vitamin D in these two kinds of cells significantly increased the expression of CYP24A1 ( P <0.001). Active vitamin D could significantly inhibit the upregulation of renin by PKA agonist forskolin, and could inhibit the expression of renin through synergy with PKA inhibitor H89., Conclusion: The expression of renin in placental decidua is up-regulated in patients with PE, and the activation of local renin-angiotensin system at the maternal-fetal interface may be involved in the pathogenesis of PE. Vitamin D can specifically down-regulate renin expression in human decidual gland epithelial cells by competing with the PKA pathway. Vitamin D supplementation may have potential value for clinical intervention of PE., (Copyright© by Editorial Board of Journal of Sichuan University (Medical Sciences).)

On May 13, 2022, World Health Organization(WHO) Position Paper on Influenza Vaccine (2022 edition) was published. This position paper updates information on influenza epidemiology, high risk population, the impact of immunization on disease, influenza vaccines and effectiveness and safety, and propose WHO's position and recommendation that all countries should consider implementing seasonal influenza vaccine immunization programmes to prepare for an influenza pandemic. In addition, it proposes that the influenza surveillance platform can be integrated with the surveillance of other respiratory viruses, such as SARS-CoV-2 and Respiratory Syncytial Virus. This position paper has some implications for the prevention and control of influenza and other respiratory infectious diseases in China: (1) Optimize influenza vaccine policies to facilitate the implementation of immunization services; (2) Influenza prevention and control should from the perspective of Population Medicine focus on the individual and community to integrate with "Promotion, Prevention, Diagnosis, Control, Treatment, Rehabilitation"; (3) Incorporate prevention and control of other respiratory infectious diseases such as influenza, COVID-19, respiratory syncytial virus and adenovirus, and intelligently monitor by integrating multi-channel data to achieve the goal of co-prevention and control of multiple diseases.

An association has been shown between chronic cigarette smoking and structural abnormalities in the brain areas related to several functions relevant to addictive behavior. However, few studies have focused on the structural alternations of chronic smoking by using magnetic resonance imaging (MRI). Also, it remains unclear how structural alternations are associated with tobacco-dependence severity and the positive/negative outcome expectances. The q-sampling imaging (GQI) is an advanced diffusion MRI technique that can reconstruct more precise and consistent images of complex oriented fibers than other methods. We aimed to use GQI to evaluate the impact of the neurological structure caused by chronic smoking. Sixty-seven chronic smokers and 43 nonsmokers underwent a MRI scan. The tobacco dependence severity and the positive/negative outcome expectancies were assessed via self-report. We used GQI with voxel-based statistical analysis (VBA) to evaluate structural brain and connectivity abnormalities. Graph theoretical analysis (GTA) and network-based statistical (NBS) analysis were also performed to identify the structural network differences among groups. Chronic smokers had smaller GM and WM volumes in the bilateral frontal lobe and bilateral frontal region. The GM/WM volumes correlated with dependence severity and outcome expectancies in the brain areas involving high-level functions. Chronic smokers had shape changes in the left hippocampal head and tail and the inferior brain stem. Poorer WM integrity in chronic smokers was found in the left middle frontal region, the right superior fronto-occipital fasciculus, the right temporal region, the left parahippocampus, the left anterior internal capsule, and the right inferior parietal region. WM integrity correlated with dependence severity and outcome expectancies in brain areas involving high-level functions. Chronic smokers had decreased local segregation and global integration among the brain regions and networks. Our results provide further evidence indicating that chronic smoking may be associated with brain structure and connectivity changes., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Neuropeptide FF (NPFF), an octapeptide of the RFamide-related peptides (FaRPs), is involved in regulatory function in various biological processes. The regulatory role of NPFF in the immune and inflammatory response was currently being revealed., Methods: Neuropeptide FF-related gene (termed LpNPFF) and its two receptors, NPFF receptor 1 (LpNPFFR1) and NPFF receptor 2 (LpNPFFR2) were identified by PCR and Semi-quantitative RT-PCR assay. Effect of LpNPFF on the production of nitric oxide (NO) in macrophage RAW264.7 cell was divided into PBS group, lipopolysaccharide (LPS) group, LPS treated with LpNPFF group, and LPS treated with LpNPFF and receptor antagonist RF9 group. Then specimens were measured by color reaction at 570 nm absorbance value., Results: Sequence analysis showed that LpNPFF cDNA consists of 835 nucleotides with a 5'- untranslated region (UTR) of 150 base pair (bp), an open reading frame (ORF) of 384 bp and a 3'-UTR of 300 bp (Accession No. MT012894). The ORF encodes 127 amino acid (aa) residues with a hydrophobic signal peptide at N-terminus and two presumptive peptides with -PQRFa structure, LpNPFF (1) and LpNPFF (2). LpNPFFR1 and LpNPFFR2 encode 427 and 444 aa residues respectively, which both have seven hydrophobic TMDs and identified as G protein coupled receptors (GPCRs). Results of tissue distribution showed that LpNPFF and receptors were highly expressed in the brain and gonad. Furtherly, in vitro assay found LpNPFF could inhibit NO production in RAW 264.7 macrophages under inflammatory stress with LPS, while its receptor antagonist RF9 caused the evoke of NO generation., Conclusions: These results contribute to the further study of neuropeptide evolution in marine organisms, and also provide a new research idea for exploring the related functions of NPFF gene., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Southwest China is home to numerous ethnic minorities, as well as many geographically and genetically isolated groups. However, the genetic substructure of these ethnic groups, especially the paternal genetic structure between groups, has not been comprehensively analyzed. In this study, we used Y chromosome capture and Illumina sequencing technologies to investigate the paternal genetic structure of three isolated groups of male unrelated individuals, including Baima in Pingwu, Sichuan Province, Muya in Shimian, Sichuan Province, and Kongge in Jinghong, Yunnan Province. We calculated the frequencies of related haplogroups by the fixed-point compound amplification method and direct counting method, and used the Past3.0 software to perform principal component analysis to draw a population clustering tree. we observed that Kongge had 3 Y chromosome haplogroups, Baima had 4 Y chromosome haplogroups, and Muya had 5 Y chromosome haplogroups. The results showed that Kongge was most closely related to the Wa, and the Y chromosome types of the Baima and Muya were mainly concentrated in the D haplogroup and its lower reaches. It has the closest relationship with the Tibetans in Qamdo and Nyingchi. The study on the genetic structure of different ethnic groups has enriched the genetic relationship of isolated populations and provided a new perspective for understanding Chinese ethnic groups.

Toxoplasmosis, caused by the intracellular protozoon Toxoplasma gondii , is a significant parasitic zoonosis with a world-wide distribution. As a main transmission route, human infection can be acquired by the ingestion of T. gondii oocysts from the environment (e.g., soil, water, fruits and vegetables). Regarding the detection of T. gondii oocysts in environmental samples, the development of a time-saving, cost-effective and highly sensitive technique is crucial for the surveillance, prevention and control of toxoplasmosis. In this study, we developed a new method by combining recombinase-aided amplification (RAA) with CRISPR-Cas12a, designated as the RAA-Cas12a-Tg system. Here, we compared this system targeting the 529 bp repeat element (529 bp-RE) with the routine PCR targeting both 529 bp-RE and ITS-1 gene, respectively, to assess its ability to detect T. gondii oocysts in soil samples. Our results indicated that the 529 bp RE-based RAA-Cas12a-Tg system was able to detect T. gondii successfully in nearly an hour at body temperature and was more sensitive than the routine PCR assay. The sensitivity of this system reached as low as 1 fM with high specificity. Thus, RAA-Cas12a-Tg system provided a rapid, sensitive and easily operable method for point-of-care detection of T. gondii oocysts in soil, which will facilitate the control of T. gondii infection in humans and animals.

Several researches reported that piscidin members of teleosts owned strong antiparasitic activity. Cryptocaryon irritans, a type of ectoparasite, could infect most of the marine teleosts. Larimichthys crocea could severely suffer from marine white spot disease caused by C. irritans, and their mortality rate was significantly high. Concentrating on this problem, we have done many related works. Piscidin 5 like (termed Lc-P5L) was another piscidin member isolated from a comparative transcriptome of C. irritans-immuned L. crocea. In the paper, quantitative Real-time PCR (qRT-PCR) showed Lc-P5L was upregulated in examined tissues, including gill, head kidney, muscle, liver, spleen and intestine after challenged by C. irritans, the significant upregulation time was in accordance to key developmental stages of C. irritans, which implied different infection stages could result in host immune response. Furthermore, using microscope techniques, we observed theronts or trophonts became weakly motile, cilia became detached, cells were out of shape, membranes eventually lysed in different cell positions and cytoplasmic contents leaked. Laser confocal scanning microscope (LCSM) observed theronts macronucleus grew swell and depolymerized after treated by recombinant Lc-P5L (rLc-P5L). Data suggested rLc-P5L was significantly lethal to C. irritans, and the death state of the parasite incubated with rLc-P5L was remarkably similar to other piscidin members or other antiparasitic peptides (APPs). Thus, these data provided new insights into L. crocea immunity against C. irritans and potential of rLc-P5L as a therapeutic agent against pathogen invasion., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

This study investigated the potential efficacy of pirarubicin (THP) in modulating rabbit conjunctival fibrosis both in vitro and in vivo and characterized the underlying mechanisms. Primary rabbit conjunctival fibroblasts (RCF) were cultured and treated with THP or mitomycin C (MMC) for 5 min, followed by assaying for cell viability, cell cycle distribution, apoptotic and autophagic pathways. The production of reactive oxygen species (ROS) and chemotaxis of macrophages by RCF were evaluated using 2',7'-dichlorofluorescein diacetate (DCFH-DA) labeling and transwell migration assay, respectively. Limbal stem cell excision in combination with alkali burn was performed on the rabbits to establish a model of limbal deficiency and conjunctival fibro-vascular invasion. After three months, the modeled fibro-vascular tissue was excised combined with topical subconjunctival 5-min exposure to THP compared with MMC intraoperatively. The recurrence of postoperative fibrosis and the expression of apoptosis, autophagy, and inflammation markers were evaluated by immunohistochemistry. All modeled rabbits developed conjunctival fibro-vascular lesions, which were similar to human recurrent pterygium (HRP). Both THP and MMC inhibited RCF proliferation and arrested cell cycle at the G0/G1 phase. In particular, 7.5 μmol/L THP remarkably promoted RCF autophagy by upregulating the levels of Beclin 1, Atg 5/12 conjugate, and LC3B, whereas, 15 μmol/L THP significantly triggered a cascade of mitochondrial-associated RCF apoptosis. THP induced the production of ROS and enhanced the chemoattraction of macrophages by RCF. Similar to 600 μmol/L MMC, both 7.5 μmol/L and 15 μmol/L THP attenuated postoperative conjunctival fibrosis in the models; 7.5 μmol/L THP preferentially enhanced autophagy while causing fewer side effects. THP exerted its antifibrotic action by modulating autophagy in RCF, inducing cell cycle arrest, and mitochondrial-mediated apoptosis. THP at the dose of 7.5 μmol/L prevented postoperative conjunctival fibrosis in an animal model., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Neuropeptides are released by neurons that are involved in a wide range of brain functions, such as food intake, metabolism, reproduction, and learning and memory. A full-length cDNA sequence of an FMRFamide gene isolated from the cuttlefish Sepia pharaonis (designated as SpFMRFamide ) was cloned. The predicted precursor protein contains one putative signal peptide and four FMRFamide-related peptides. Multiple amino acid and nucleotide sequence alignments showed that it shares 97% similarity with the precursor FMRFamides of Sepiella japonica and Sepia officinalis and shares 93% and 92% similarity with the SpFMRFamide gene of the two cuttlefish species, respectively. Moreover, the phylogenetic analysis also suggested that Sp FMRFamide and FMRFamides from S. japonica and S. officinalis belong to the same sub-branch. Tissue expression analysis confirmed that SpFMRFamide was widely distributed among tissues and predominantly expressed in the brain at the three development stages. The combined effects of Sp FMRFamide+ Sp GnRH and Sp FLRFamide+ Sp GnRH showed a marked decrease in the level of the total proteins released in the CHO-K1 cells. This is the first report of SpFMRFamide in S. pharaonis and the results may contribute to future studies of neuropeptide evolution or may prove useful for the development of aquaculture methods for this cuttlefish species.

Antibiotic resistance genes (ARGs) in municipal wastewater pose a potential threat to the environment. In this study, the change in absolute and relative abundance of ARGs, metal resistance genes (MRGs), and mobile genetic elements (MGTs) were investigated during an emergent municipal wastewater treatment by the magnetic separation process. Results indicate that all the concentrations of targeted ARGs, MRGs, and MGTs decreased significantly in the primary and secondary stirring tank. However, the absolute abundance of some ARGs and MRGs increased in the effluent, which is likely caused by the presence of ample MGTs, in the order of int1 (2.00×10 10 copies·mL -1 ) > int2 (1.91×10 8 copies·mL -1 ) > Tn 916/1545 e (5.38×10 8 copies·mL -1 ). The results obtained from network and PCA analysis showed that the removal of ARGs and MRGs were significantly associated with variations in the microbial community and common pollutants in urban wastewater, such as suspended solids, phosphorus, and COD, which are important factors for affecting the removal efficiency of antibiotic resistance genes and metal heavy resistance genes. These results show that magnetic separation can effectively reduce common pollutants in urban wastewater and might further restrict the transmission and transfer of ARGs. Moreover, it is necessary to strengthen the subsequent management of magnetic separation effluent and dehydrated sludge by disinfection technologies to lessen the risk of antimicrobial contamination.

Chemical drugs, such as antibiotics, were still important materials to prevent and cure diseases of aquatic organisms. However, antibiotics abuse do not only make the effects little, but also cause other bad problems, such as bacterial resistance and drug residues. Therefore, seeking the effective substitutes of antibiotics was an approach needed to be explored. Antibacterial peptides (AMPs) attracted more and more attention in the recent years. The parasitism and secondary bacterial invasion caused by ectroparasite Cryptocaryon irritans was a disaster to almost all host fish, including Larimichthys crocea. Reports indicated many AMPs played a key role in the whole parasitic infection cycle. Piscidin 5 like was a member of piscidin family. In the study, the antibacterial activity and mechanisms of piscidin 5 like from L.coreca (Lc-P5L) were detected. Liquid growth inhibition results showed recombinant Lc-P5L (rLc-P5L) had broad antibacterial spectrum and strong bactericidal activity. The bactericidal activity functioned in dose- and time-dependent manners. SEM (scanning electron microscope) observed the relatively detailed bactericidal process, rLc-P5L treatment resulted in a mass of bacteria piling together, appearing plenty of strange filaments and covering on the bacteria. Besides, S.aureus overgrowed plenty of granules, formed holes on the membrane of a few cells, and contents poured out from the holes. At the same time, antibacterial mechanisms were explored. After direct incubation with bacteria, western blot detected the apparently positive signal of rLc-P5L on bacteria; secondly, the incubation first with LPS (lipopolysaccharide) or LTA (lipoteichoic acid) significantly affect the binding of rLc-P5L to bacteria again, which indicated rLc-P5L could bind to bacteria through interaction with some PAMPs (pathogen-associated molecular patterns). In addition, rLc-P5L could interact with bacterial genome DNA by dose- and time-dependent means. In summary, rLc-P5L binded to bacteria surface through targeting to some PAMPs to damage membrane, and entered into cells to interact with genome DNA to disturb normal metabolism when it reached to some certain time and concentration thresholds, which were likely to be its pathway to exert antibacterial activity., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Objective: To explore the effects of macrophages with high expression of TL1A on the activation and proliferation of HSCs in vitro. Methods: The Bone marrow-derived macrophages (BMMs) and peritoneal macrophages (PMs) from wild type (WT) and myeloid-overexpressed TL1A transgenic mice were isolated, differentiated and activated. HSCs were harvested from activated macrophages culture supernatant (CM). HSCs were detected by immunofluorescence and real-time Q-PCR. And the proliferation was detected by CCK-8 and BrdU assay kit. The levels of IL-1β and PDGF-BB in macrophage culture supernatants were determined by enzyme-linked immunosorbent assay (ELISA). Results: BMMs-derived CM-intervention HSCs were used to detect the expression of α-smooth muscle actin (α-SMA) on the 2nd, 4th and 6th day respectively by immunofluorescence method. There was no significant difference between the two groups on the 2 nd and the 6th day, P > 0.05; On day 4, the CM/Tg group was significantly higher than that of CM/WT group, P < 0.01; the results of CMs derived from PMs were consistent with the above trend. The expression of α-SMA mRNA on the 2nd, 4th and 6th day was detected by real-time Q-PCR method using BM-derived CMs. No significant difference was found between the groups on the 2nd day ( P > 0.05).α-SMA mRNA increased further on the 4th and 6th day, and the level of CM/Tg in CM/Tg group was significantly higher than that in CM/WT group ( P < 0.05). The detection results of CMs derived from PMs were consistent with the above trend. The results of CCK-8 assay and BrdU assay showed that the proliferation rate of HSCs in CM Tg group was significantly higher than that in CM/WT group ( P < 0.01). The CMs derived from PMs were used to interfere with HSCs. And the results were consistent with the above trend. For BMMs, the levels of IL-1β and PDGF-BB in the lipopolysaccharide (LPS) + IFNγ/Tg culture supernatant were significantly higher than those in the LPS+IFNγ/WT group ( P < 0.01). For the culture supernatants of PMs Liquid test results consistent with the above trend. Conclusion: Macrophages with high expression of TL1A could enhance the activation and proliferation of HSCs by increasing the secretion of IL-1β and PDGF-BB.

Larimichthys crocea, the special marine economy fish, owns the largest annual yield for a single species in China. One of the most significant factors affecting large yellow croaker culture is the diseases, especially the threat of marine white spot disease which caused by a protozoan Cryptocaryon irritans. Antimicrobial peptides (AMPs) have been demonstrated to be active against bacterium, fungi and parasites, showing their potential usefulness in aquaculture as substitutes for antibiotics. Many researches have been carried out about the AMPs concentrating on the activity resist on C. irritans, and piscidin-like of L. crocea owning widely antibacterial spectrum and strong activity against C. irritans was screened in our team. In the paper, taking advantage of the large yellow croaker hepatic comparison transcriptome in response to C. irritans at 3d post infection, seven kinds of AMPs have been excavated from the differently expressed genes, including LEAP2 like, LEAP-2A, hepcidin, hepcidin-like, piscidin-5-like, piscidin-5-like type 4 and bactericidal permeability increasing protein (BPI). Hepcidin, hepcidin-like, piscidin-5-like, piscidin-5-like type4 and BPI were up-regulated to protect large yellow croaker from being damaged by C. irritans infection; while LEAP2 like and LEAP-2A were down-regulated, they might be as a negative-feedback regulation factor or some other regulatory mechanisms to adjust the immune response in the process of C. irritans infection. The differential expression changes were verified with quantitative real-time PCR (qRT-PCR) to illustrate the reliability of the sequenced data. Hearteningly, piscidin-5-like type 4 was a novel type which was high similar to other piscidin-5-like types. Interestingly, the infection may well cause alternative splicing of LEAP-2A mRNA, which was a surprised phenomenon and finding after C. irritans infection, but more further study was needed to be conducted. Therefore, the data showed that these AMPs were involved in the immune response to the C. irritans infection. In all, these results implied that the immune response of AMPs to C. irritans infection was a complex and sophisticated regulatory process., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Objective: To investigate the possibility of the visual system homeobox 1 (VSX1) gene as a candidate susceptibility gene for Chinese patients with sporadic keratoconus, and to identify sequence variants of the VSX1 gene in such patients. Methods: Cross-sectional study. Genomic DNA was extracted from the leukocytes in the peripheral venous blood of 50 patients with sporadic keratoconus and 50 control subjects without this ocular disorder. Five exons and the intron-exon splicing of the VSX1 gene were amplified by polymerase chain reaction (PCR). The PCR products were directly sequenced and compared to the GeneBank database to find mutations. Bioinformatics analysis was done to predict the influence of these mutations on proteins. Results: One novel missense heterozygous mutation (p.R131P) was found in exon 1 of the VSX1 gene in one keratoconus patient. Another heterozygous mutation (p.G160V) in exon 2 was found in two keratoconus patients. These mutations were not detected in the control subjects. Bioinformatics analysis predicted that the p.R131P mutation may not cause a pathogenic change, but the p.G160V mutation might be functionally deleterious. In intron 3 of the VSX1 gene, the nucleotide substitution of g.8326G>A was detected to be heterozygous in 3 cases of sporadic keratoconus and 4 cases of control and homozygous in 2 cases of sporadic keratoconus and 1 case of control. The variation of g.8326G>A belonged to a single polymorphism change of the VSX1 gene. Conclusions: The p.R131P detected in this study is a novel mutation of the VSX1 gene. Sequence variants of the VSX1 gene were identified for the first time in Chinese patients with sporadic keratoconus, but their precise role in disease causation requires further investigations. (Chin J Ophthalmol, 2018, 54: 212-217) .

Aim: To investigate the hepatic differentiation potential of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) and to evaluate their therapeutic effect on liver fibrosis/cirrhosis., Methods: A CCl 4 -induced liver fibrotic/cirrhotic rat model was used to assess the effect of hUC-MSCs. Histopathology was assessed by hematoxylin and eosin (H&E), Masson trichrome and Sirius red staining. The liver biochemical profile was measured using a Beckman Coulter analyzer. Expression analysis was performed using immunofluorescent staining, immunohistochemistry, Western blot, and real-time PCR., Results: We demonstrated that the infused hUC-MSCs could differentiate into hepatocytes in vivo . Functionally, the transplantation of hUC-MSCs to CCl 4 -treated rats improved liver transaminases and synthetic function, reduced liver histopathology and reversed hepatobiliary fibrosis. The reversal of hepatobiliary fibrosis was likely due to the reduced activation state of hepatic stellate cells, decreased collagen deposition, and enhanced extracellular matrix remodeling via the up-regulation of MMP-13 and down-regulation of TIMP-1., Conclusion: Transplanted hUC-MSCs could differentiate into functional hepatocytes that improved both the biochemical and histopathologic changes in a CCl 4 -induced rat liver fibrosis model. hUC-MSCs may offer therapeutic opportunities for treating hepatobiliary diseases, including cirrhosis., Competing Interests: Conflict-of-interest statement: We declare that there are no conflicts of interest to disclose.

The marine white spot disease caused by protozoan ectoparasite Cryptocaryon irritans is a severe problem to the large yellow croaker farming industry. To understand the molecular immune mechanisms and improve its immune capacity are particular important. STING, one of the important second messengers in innate immune response process, plays pivotal roles in defensing against different pathogenic microorganisms. Many reports have pointed that STING could not only combine the uncovered dsDNA, ssDNA directly in the cytoplasm from the pathogens or biology itself, but it also could recognize cyclic diguanylate monophosphate (c-di-GMP), cyclic diadenylate monophosphate (c-di-AMP). Based on the STING sequence, a variant of the L. crocea STING (termed LcSTING2) was found by accident. RACE was used to clone the full-length cDNA of LcSTING2 which contained a 5'- UTR of 154 bp, a 3'-UTR of 592 bp and an ORF of 1227 bp encoding 408 amino acids. The predicted protein molecular weight (Mw) was 45.83 KDa and the estimated theoretical isoelectric point (pI) was 6.24. The deduced protein of LcSTING2 contains no signal peptide. One transmembrane motif (TM) in the N-terminal region, a TMEM173 domain and five putative motifs (RXR) found in resident endoplasmic reticulum proteins were also conserved. According to the partial genomic sequence, the unknown sequences were amplified, it contained 6 exons and 5 introns, and all the exon-intron boundaries were in accordance with classical GT-AG rule (GT/intron/AG). The similarity shared with fishes was higher than other high vertebrates. qRT-PCR results showed that LcSTING (two variants) distributed in all examined tissues, and it was the most abundant in gill. After challenged by C. irritans, LcSTING mRNA only appeared instantaneous up-regulation during the infective stage of theronts in the head kidney and was also up-regulated during the whole infectious cycle of C. irritans in the liver, which implied LcSTING gene was likely to be involved in the defensing against C. irritans infection, it is the first time to explore the STING taking part in the immune response to ectoparasite rather than bacterium or viruses., (Copyright © 2017. Published by Elsevier Ltd.)

Background: Keratoconus normally presents as a sporadic disease. Although different studies have found sequence variants of the visual system homeobox 1 (VSX1) gene associated with keratoconus in humans, no research has detected such variants in sporadic keratoconus patients from China. To investigate the possibility of VSX1 being a candidate susceptibility gene for Chinese patients with sporadic keratoconus, we performed sequence screening of this gene in such patients., Methods: Whole DNA was obtained from the leukocytes in the peripheral venous blood of 50 patients with sporadic keratoconus and 50 control subjects without this ocular disorder. Polymerase chain reaction single-strand conformation polymorphism analysis and direct DNA sequencing technology were used to detect sequence variation in the five exons and splicing regions of the introns of the VSX1 gene. The sequencing results were analyzed using DNAstar software., Results: One novel missense heterozygous sequence variant (p.Arg131Pro) was found in the first exon of the VSX1 gene in one keratoconus patient. Another heterozygous sequence variant (p.Gly160Val) in the second exon was found in two keratoconus patients. These variants were not detected in the control subjects. In the third intron of the VSX1 gene, c.8326G > A nucleotide substitution (including heterozygous and homozygous change) was also discovered. The frequency of this variation did not differ significantly between patients and controls, it should belong to single-nucleotide polymorphism of the VSX1 gene. Bioinformatic analysis also predicted that one missense sequence variation (p.Arg131Pro) may not cause a pathogenic change., Conclusions: In this study, we added one novel missense sequence variation (p.Arg131Pro) in the coding region of the VSX1 gene to the range of VSX1 coding region variations observed in patients with sporadic keratoconus from China. Our work suggests that VSX1 sequence variants might be involved in the pathogenesis of sporadic keratoconus, but their precise role in disease causation requires further investigation.

Aim: To evaluate the effects of phosphatase and tension homologue deleted on chromosome ten ( PTEN ) gene on collagen metabolism in hepatic fibrosis and the underlying mechanisms., Methods: Rat primary hepatic stellate cells (HSCs) and human LX-2 cells were transfected with adenovirus containing cDNA constructs encoding wild-type PTEN (Ad-PTEN), PTEN mutant G129E gene (Ad-G129E), and RNA interference constructs targeting the PTEN sequence PTEN short hairpin RNA to up-regulate and down-regulate the expression of PTEN . HSCs were assayed using fluorescent microscopy, real-time polymerase chain reaction, and western blotting. Moreover, a CCl 4 -induced rat hepatic fibrosis model was established to investigate the in vivo effects. Hematoxylin and eosin, and Masson's trichrome were used to assess the histological changes. The expression of collagen I and III was assessed using immunohistochemistry and western blot analysis., Results: Elevated expression of PTEN gene reduced serum levels of alanine transaminase and aspartate transaminase, decreased collagen deposition in the liver, and reduced hepatocyte necrosis. In contrast, knockdown of PTEN expression had an opposite effect, such as increased collagen deposition in the liver, and was molecularly characterized by the increased expression of matrix metalloproteinase (MMP)-13 ( P < 0.01) and MMP-2 ( P < 0.01), as well as decreased expression of the tissue inhibitor of metalloproteinase (TIMP)-1 ( P < 0.01) and TIMP-2 ( P < 0.01)., Conclusion: These data indicated that gene therapy using recombinant adenovirus encoding PTEN might be a novel way of treating hepatic fibrosis., Competing Interests: Conflict-of-interest statement: We declare that there are no conflicts of interest to disclose.

Background: Reis-Bücklers corneal dystrophy (RBCD) was consistently reported as a corneal dystrophy only affected Bowman's layer and superficial corneal stroma, and superficial keratectomy was a recommendation surgery for treatment in literatures. The study reported new histopathological and ultrastructural findings in RBCD caused by the Arg124Leu mutation of transforming growth factor induced (TGFBI) gene in a four-generation Chinese pedigree., Methods: Subjects including eight patients and seven unaffected family members received slit-lamp biomicroscopy and photography. DNA was obtained from all subjects, and exons 4 and 11 to 14 of TGFBI gene were analyzed by polymerase chain reaction and the products were sequenced. Anterior segment optical coherence tomography (AS OCT) and in vivo confocal microscopy were conducted for ten eyes of five patients. Based on the results of AS OCT and in vivo confocal microscopy, deep anterior lamellar keratoplasty (DLKP) using cryopreserved donor cornea was applied for four eyes of four patients. Four lamellar dystrophic corneal buttons were studied by light and transmission electron microscopy, and TGFBI immunohistochemistry., Results: Eight patients had typical clinical manifestations of RBCD presenting recurrent painful corneal erosion starting in their early first decades, along with age-dependent progressive geographic corneal opacities. TGFBI sequencing revealed a heterozygous mutation, Arg124Leu in all eight patients. Anterior segment optical coherence tomography and in vivo confocal microscopy showed the dystrophic deposits involved not only in subepithelial and superficial stroma, but also in mid- or posterior stroma in four examined advanced eyes. Light microscopy showed Bowman's layer was absent, replaced by abnormal deposits stain bright red with Masson's trichrome. In superficial cornea, the deposits stacked and produced three to five continuous bands parallel to the corneal collagen lamellae. In mid- to posterior stroma, numerous granular or dot- like aggregates were heavily scattered, and most of them presented around the nuclei of stromal keratocytes. Transmission electron microscopy revealed the multiple electron-dense rod-shaped deposits aggregated and formed a characteristic pattern of three to five continuous bands in superficial cornea, which were similar to those seen under light microscopy. In mid- to posterior stroma, clusters of rod-shaped bodies were scattered extracellular or intracellular of the stromal keratocytes between the stromal lamellae suggesting the close relationship between mutated proteins and keratocyte., Conclusions: The study offer evidences indicating DLKP is a viable treatment option for advanced RBCD to avoid recurrence, and the mutated TGFBIp in dystrophic corneas are of keratocytes origin.

White spot syndrome virus (WSSV) is a lethal pathogen of shrimp and many other crustaceans, including crayfish. However, the molecular mechanism underlying its cellular entry remains elusive due to the lack of shrimp cell lines for viral propagation. Crayfish hematopoietic tissue (Hpt) cell culture was recently established as a good model for WSSV infection study. Here, we showed that multiple endocytic routes, including clathrin-mediated endocytosis (CME), macropinocytosis and caveolae-mediated endocytosis, were indispensably employed for the viral entry into Hpt cell of the crayfish Cherax quadricarinatus. Intriguingly, cellular autophagic activity was positively correlated with efficient viral entry, in which a key autophagy-related protein, γ-aminobutyric acid receptor-associated protein (Cq-GABARAP), that not only localized but also co-localized with WSSV on the Hpt cell membrane, strongly facilitated WSSV entry by binding to the viral envelope VP28 in a CME-dependent manner that was negatively regulated by Cq-Rac1. Furthermore, cytoskeletal components, including Cq-β-tubulin and Cq-β-actin, bound to both recombinant rCq-GABARAP and WSSV envelope proteins, which likely led to viral entry promotion via cooperation with rCq-GABARAP. Even under conditions that promoted viral entry, rCq-GABARAP significantly reduced viral replication at an early stage of infection, which was probably caused by the formation of WSSV aggregates in the cytoplasm.

Objective: This study aims to explore the ocular surface of late stage eye burns by impression cytology (IC) and analyze the cytological changes and their relationship to ocular surface abnormalities., Patients and Methods: 68 eyes with late stage eye burns (thermal burn: 28 eyes; alkali burn: 26 eyes; acid burn: 14 eyes), procured from 68 patients (aged ranges from 17 to 70 years old). Ocular surface abnormalities were assessed under slit lamp and graded. These were broadly classified as eyelid, corneal, conjunctival, and tear film abnormalities. Impression cytological examination was taken by cellulose acetate filter paper for all eyes. Samples were analyzed and scored under light microscope, including the status of epithelial cells, goblet cells, mucus and inflammatory cells. All the results and data were compared and analyzed by SPSS software (version 16.0)., Results: According to the IC results, loosed cell-to-cell density and nuclear abnormality, keratinization, reduced goblet cell amount, disorder of mucus, and existing of inflammatory cells were observed in almost all the cases. The IC results were significantly correlated to the ocular surface injury severity (r=0.458, p<0.01). The ocular surface injury severity mostly contains three aspects: the corneal neovascularization scales, the present or absent of recurrent epithelial erosion and the tear film break-up time. Eyes with the foreword three symptoms were inclined to have higher IC scores. The epithelial cell-to-cell density, goblet cell and mucus amount were all correlated to tear film break-up time. However, inflammatory cell density showed no significant correlation to the conjunctival hyperemia grade. But inflammatory cell density correlated to the corneal opacity grade and epithelial stability status., Conclusions: IC examinations could reflect the cytological disorders and relative injury severity of the ocular surface in late stage eye burns. It provides further information which will be useful in surgery and therapy.

Objective: This study aims to investigate the effects of sudden load changes (expected and unexpected imbalance) on the activity of muscles of the lumbar spine and their central motor control strategy in military personnel with or without chronic low back pain (LBP)., Design: Bilateral sudden imbalance was examined (2 × 2 factorial design)., Setting: The 117th PLA Hospital, Hangzhou, China, Participants: Twenty-one male subjects with lower back pain and 21 male healthy control subjects were active members of the Nanjing Military Region land forces., Outcome Measures: Independent variables: LBP vs. healthy controls and imbalance anticipation (expected and unexpected imbalance)., Dependent Variables: rapid reaction time (RRT) and intensity of rapid reaction (IRR) of bilateral lumbar (L3-L4) erector spinae (ES), lumbar (L5-S1) multifidus (MF), and abdominal external oblique muscles. Results Under expected or unexpected sudden imbalance conditions, subjects with LBP demonstrated significantly greater IRR than healthy controls in ipsilateral and contralateral ES and MF, respectively (P < 0.05 for all). IRR of contralateral ES was significantly larger than that of the ipsilateral ES. A significant group effect of RRT of both ipsilateral and contralateral ES muscles and a significant time expectation effect on RRT of contralateral MF muscles were also observed. RRT of the contralateral ES muscles was significantly lower than that of the ipsilateral ES muscles (P < 0.001)., Conclusions: Sudden imbalance prolonged RRT of selected trunk muscles in patients with chronic LBP. The activation amplitude increased. The results may provide a theoretical basis for a study on the pathogenesis of chronic LBP.

Inflammatory bowel disease (IBD) results from a complex series of interactions between susceptibility genes, the environment, and the immune system. The host microbiome, as well as viruses and fungi, play important roles in the development of IBD either by causing inflammation directly or indirectly through an altered immune system. New technologies have allowed researchers to be able to quantify the various components of the microbiome, which will allow for future developments in the etiology of IBD. Various components of the mucosal immune system are implicated in the pathogenesis of IBD and include intestinal epithelial cells, innate lymphoid cells, cells of the innate (macrophages/monocytes, neutrophils, and dendritic cells) and adaptive (T-cells and B-cells) immune system, and their secreted mediators (cytokines and chemokines). Either a mucosal susceptibility or defect in sampling of gut luminal antigen, possibly through the process of autophagy, leads to activation of innate immune response that may be mediated by enhanced toll-like receptor activity. The antigen presenting cells then mediate the differentiation of naïve T-cells into effector T helper (Th) cells, including Th1, Th2, and Th17, which alter gut homeostasis and lead to IBD. In this review, the effects of these components in the immunopathogenesis of IBD will be discussed.

Gracilaria eucheumoides Linn (Gracilariaceae; G. eucheumoides ) is abundant in dietary fiber, which aids the clearance of excess cholesterol from the blood and maintains stable blood glucose levels. The aim of the present study was to investigate the antifatigue effect of G. eucheumoides in mice and the physiological and molecular mechanisms underlying this effect. Mice were randomly divided into four groups and three of the groups were administered different doses of G. eucheumoides extract. A loaded swimming test demonstrated that the swimming times of the low-, medium- and high-dose groups were longer than those of the control group. Examinations revealed that the liver and muscle glycogen, lactate dehydrogenase and blood glucose concentration levels of the treatment groups were higher than those of the control group (P<0.05). However, this was not the case for lactic acid concentration (P>0.05). Quantitative polymerase chain reaction showed that the gene expression levels of glucose transport protein 4 and AMP-activated protein kinase in the medium-dose group exhibited the largest increases, compared with the other treatment groups, and were 3.0- and 1.8-fold higher than those in the control group, respectively. The results of the present study indicated that G. eucheumoides exerts an antifatigue effect on mice.

Background: Diabetic patients have a high prevalence of shoulder pain and stiffness. Interleukin (IL) 1β was reportedly correlated with shoulder stiffness and decreased shoulder function. We retrospectively compared the expression of IL-1β in the subacromial synovial fluid between diabetic and nondiabetic patients with rotator cuff tearing., Materials and Methods: We enrolled 68 patients with rotator cuff tearing (23 diabetic patients and 45 nondiabetic patients). The preoperative sum of range-of-motion deficit (SROMD), Constant score, and visual analog scale (VAS) score were obtained. Intraoperatively, subacromial synovial fluid was collected for the IL-1β level measurement. Comparisons of IL-1β levels, Constant scores, SROMD, and VAS scores between diabetic and nondiabetic patients were analyzed with the Mann-Whitney U test., Result: Diabetic patients with rotator cuff tearing had significantly increased subacromial IL-1β levels (P = .048), SROMD (P < .001) and VAS scores (P = .022) and lower Constant scores (P < .001) than nondiabetic patients. The IL-1β levels in the subacromial fluid were significantly correlated with the Constant score (r = -0.477, P < .001), VAS score (r = 0.698, P < .001), and SROMD (r = 0.293, P = .015) in all patients., Conclusion: The elevated IL-1β levels in the subacromial fluid of patients with diabetes may explain the likelihood of pain and shoulder stiffness developing in these patients. We suggest more aggressive treatment for rotator cuff lesions in diabetic patients., (Copyright © 2013 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Mosby, Inc. All rights reserved.)

Soil samples were collected from 18 sites at a relocated lead accumulator factory in the Southwest region, China. Among the 15 sample sites, profile soil samples at 0-20 cm, 20-40 cm and 40-60 cm were taken. Soil lead contents were analyzed by inductively coupled plasma-optical emission spectroscopy (ICP-OES). Characteristics of Pb in topsoil and soil profiles collected from different sampling sites were discussed. Results showed that: (1) The total Pb contents in topsoil (0-20 cm) ranged from 18.18 to 52,332.50 mg x kg(-1). The maximum content greatly exceeded the national standard (HJ 350-2007). The Pb concentration in different workshops followed a decreasing order: the fourth workshop > the second workshop > waste lead storage pit > sewage works > the third workshop > the fifth workshop > the first workshop > the original fourth workshop > the packing workshop > the office area. (2) Results of profile distribution showed that soil depth had no significant effect on Pb content. Lead can be highly accumulated at different depths of the soil, which was quite different from natural soils. (3) Hakanson's potential ecological risk index evaluation showed that there was widespread ecological risk in the soil of the plant, and serious ecological risk existed in some workshops where a large number of lead was enriched. Therefore, the site must be restored in order to reuse it.

Professor LIN Lin's clinical experiences of comprehensive internal and external therapy for asthma was summarized in the paper. According to professor LIN Lin's experiences, asthma was differentiated as cold, heat, wind, stasis and deficiency pattern/syndrome in clinic. Asthma of cold type was treated with Xiao qinglong Decoction and Sanzi Yangqin Decoction combined with cupping, moxibustion, acupuncture and Bian stone therapy. Asthma of heat type was treated with Dingchuan Decoction combined with acupuncture, scraping therapy and moving cupping. Asthma of wind type was treated with Masu Erchen Decoction combined with acupuncture and cupping. Asthma of stasis type was treated with Xue fu Zhuyu Decoction combined with acupuncture, tapping manipulation, moving cupping and Bian stone therapy. Asthma of deficiency type was treated with Buzhong Yiqi Decoction, Shenge Shenqi Pill combined with warming needling therapy, acupoint injection and Bian stone therapy. A medical case was enclosed to explain the diagnosis and treatment idea "seeking the root cause of the disease in treatment" and "strengthening the antipathogenic qi to eliminate pathogenic factors" in the comprehensive internal and external therapy.

Background: Low back pain (LBP) is a major medical and social problem among working populations and is associated with high medical expense, loss of productivity, and disability. The aim of this study is to investigate the prevalence of LBP among soldiers and evaluate the possible causative factors in military training. The results may provide an insight into changes needed in military training that will reduce the occurrence of LBP among soldiers., Methods: A cross-sectional survey was conducted in a group of young soldiers in China to estimate the prevalence of LBP and evaluate possible causative factors in military training., Results: The survey was distributed to 1659 soldiers, of whom 1624 responded. LBP was reported by 425 of the 1624 (26.2%) soldiers. The prevalence of LBP was higher in the armored force (51.3%) than in the artillery (27.5%) or infantry (11.9%). A multivariate logical regression analysis identified night training, 5 km cross-country race, and grenade-throwing training as military training risk factors for LBP., Conclusions: The relatively high incidence of LBP among soldiers was related to night training, 5 km racing, and grenade throwing. Modifications in these training methods should enhance the health of recruits and lower the incidence of LBP.

Objective: To investigate the age variations of head-face morphological traits of Hakka in Guangdong Province., Methods: Thirty-eight physical indices of head and face of 671 Hakka adults (151 urban males, 181 urban females, 162 rural males and 177 rural females) from Meizhou city of Guangdong were investigated. Twelve physical indices of head and face were calculated and counted its indices distributions. The age changes of morphological traits of head and face were analyzed., Results: With the age growing, the rate of eyefold of the upper eyelid was increased, and the Mongoloid fold decreased, the direction of eyeslits tended to level, nasal root height dropped, the zygomatic projection was more projected, the eye color turned to lighter, the upper red lips became thinner; medium-sized facial height, nasal height, upper lip skin height, physiognomic ear length, physiognomic ear breadth and breadth of alae nasi were decreased. The index of mouth breadth, morphological facial skin fold were positively correlated with the age. The index of minimum frontal breadth, face breadth, bigonial diameter, interocular breadth, external biocular breadth, lip height, thickness of lips and horizontal head circumference were negatively correlated with the age. With the age growing, morphological facial index and vertical cephalo-facial index was ascending. Moreover, length-height index of head, transverse frontoparietal index, zygomatico-frontal index and lip index were declining., Conclusion: With the age growing, the head and face measurements, physical indices and the proportion of observed indices have changed in Hakka in Guangdong province.

Matrix metalloproteinase 9 (MMP9) and MMP2 are important in the development and maintenance of neuropathic pain behavior induced by peripheral nerve injury. The enzymatic activity of MMP9 and MMP2 is balanced specifically by tissue inhibitor of metalloproteinase 1 (TIMP1) and TIMP2, respectively. In present study, we measured the effect of peripheral nerve injury on the expression of TIMP1 and TIMP2 in adult dorsal root ganglia (DRG). A dramatic increase of TIMP1 mRNA and a decrease of TIMP2 in DRG after sciatic nerve transection (SNT) were displayed through a real-time PCR method. Furthermore, data showed by in situ hybridization that TIMP1 mRNA was only localized in DRG satellite cells under normal conditions. TIMP1 mRNA was increased in satellite cells, and induced within sensory neurons after SNT. Analysis of neuronal profiles showed that induced TIMP1 mRNA was mainly contained in small and medium DRG neurons. Further study displayed that induced TIMP1 mRNA was predominantly present in activating transcription factor 3 (ATF3)-positive injured DRG neurons. Comparatively, TIMP2 mRNA was mostly contained within sensory neurons and the overall amount decreased at the late stage after nerve injury. These data showed different change of TIMPs in DRG after peripheral nerve injury., (Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.)

Objective: To explore the association between the levels of serum growth differentiation factor-15(GDF-15) and acute coronary syndrome(ACS) in patients undergoing coronary angiography(CAG)., Methods: In this prospective study, 120 candidates (30 healthy controls and 90 patients with ACS) admitted to our hospital from March 2006 to April 2008 were included. The patients with ACS were divided into 3 groups: (1)Group AMI-emergency (AMI-E), including 30 patients with acute myocardial infarction (AMI) who underwent CAG within 12 hours of the onset of symptoms. (2) Group AMI 7-14 d, including 30 patients with AMI who underwent CAG within 7-14 days of the onset of symptoms. (3)Group UAP including 30 patients with unstable angina pectoris. The serum level of GDF-15 was determined by ELISA., Results: The level of GDF-15 for group ACS was (649.0 ± 224.21) ng/L, which was significantly higher than that of control group [(537.2 ± 262.9)ng/L,P<0.01]. The level of serum GDF-15 significantly increased in group AMI-E [(801.6 ± 218.4)ng/L] compared with group AMI 7-14 d [(656.2 ± 252.4)ng/L,P=0.042, 95% CI 1.001-1.061], group UAP [(586.9 ± 225.6)ng/L, P=0.001, 95% CI 0.809-0.950] and control group(P<0.05, 95% CI 1.000-1.012) after being adjusted for relevant covariates. The elevated level of GDF-15 was related to biomarkers, such as white blood cells count (WBC, r(2)=0.276, P=0.002), lactate dehyerogenase(LDH, r(2)=0.288, P=0.004), creatine kinase 2(CK-MB, r(2)=0.350, P<0.001) and troponin T(TnT, r(2) =0.344, P=0.001)., Conclusion: GDF-15 protein expression increases in ACS, and the level of GDF-15 rapidly increases and remains elevated in the early period of acute myocardial infarction. The elevated level of GDF-15 is related to biomarkers, such as WBC, LDH, CK-MB, and TnT. GDF-15 could be a new and independent biomarker in evaluating the patients with cardiovascular disease.

By the methods of field sampling and laboratory analysis, this paper studied the variations of soil organic carbon (SOC) and total nitrogen (TN) contents and SOC density under different land use types in Shanghai. Significant differences were observed in the test parameters among different land use types. The SOC density was the highest in paddy field (3.86 kg x m(-2)), followed by in upland (3.17 kg x m(-2)), forestland (3.15 kg x m(-2)), abandoned land (2.73 kg x m(-2)), urban lawn (2.65 kg x m(-2)), garden land (2.13 kg x m(-2)), and tidal flat (1.38 kg x m(-2)). The assessment on the effects of three types of land use change on the test parameters showed that the conversion of paddy field into upland resulted in a significant decrease of SOC and TN contents and SOC density; the abandonment of farmland was not an effective way in improving SOC storage in the Yangtze Delta region with abundant water and heat resources, high soil fertility, and high level of field management; while the 4-5 years conversion of paddy field into artificial forestland decreased the SOC and TN contents and SOC density, suggesting that in a short term, the soil carbon sequestration effect of the conversion from paddy field to forestland was at a low level, due to the limitation of vegetation productivity.

Aim: To investigate the effects and mechanism of disruption of focal adhesion kinase (FAK) expression on collagen metabolism in rat hepatic stellate cells (HSC)., Methods: The plasmids expressing FAK short hairpin RNA (shRNA) were transfected into HSC-T6 cells, and the level of FAK expression was determined by both real-time quantitative polymerase chain reaction (Q-PCR) and Western blotting analysis. The production of type I collagen and type III collagen in FAK-disrupted cells was analyzed by real-time Q-PCR. The level of collagen metabolism proteins, including matrix metalloproteinases-13 (MMP-13) and tissue inhibitors of metalloproteinases-1 (TIMP-1) was also determined by both real-time Q-PCR and Western blotting analysis., Results: The transfection of FAK shRNA plasmids into HSC resulted in disrupted FAK expression. Compared with the HK group, the levels of type I collagen and type III collagen mRNA transcripts in FAK shRNA plasmid group were significantly decreased (0.69 +/- 0.03 vs 1.96 +/- 0.15, P = 0.000; 0.59 +/- 0.07 vs 1.62 +/- 0.12, P = 0.020). The production of TIMP-1 in this cell type was also significantly reduced at both mRNA and protein levels (0.49 +/- 0.02 vs 1.72 +/- 0.10, P = 0.005; 0.76 +/- 0.08 vs 2.31 +/- 0.24, P = 0.000). However, the expression of MMP-13 mRNA could be significantly up-regulated by the transfection of FAK shRNA plasmids into HSC (1.74 +/- 0.20 vs 1.09 +/- 0.09, P = 0.000)., Conclusion: These data support the hypothesis that shRNA-mediated disruption of FAK expression could attenuate extracellular matrix (ECM) synthesis and promote ECM degradation, making FAK a potential target for novel anti-fibrosis therapies.

This paper presents a non-invasive and non-ionizing radiation method for assessment of osteoporosis by analyzing in vivo MR images. Texture features (entropy, coarse, etc...) derived from co-occurrence matrix and neighborhood graystone difference matrix are obtained. These features are significantly different between patients and control subjects. Then the grayscale MR image is transformed to the binary image, The shape and topology features (area, skeleton length, euler number, etc. ) obtained from the binary image show too significant differences between patients and control subjects.

From 1998 to 2001, a sample of 2760 Mongol and Han individuals was investigated in 3 areas (Eerduosi city, Xilinguole and Bayannaoer leagues) from Inner Mongolia. Each individual was studied on 4 genetic characters (thumb type, palmar digital formular, fingernail type and plantar digital formular). The result is as follows: (1) The frequencies of the straight thumb were over 60%, and there were no significant sexual differences in all of 6 groups. (2) The frequencies of the ring-finger-longer were much higher in 6 groups, and there were significant sexual differences in 4 groups. (3) The frequencies of the long-shaped fingernail were the highest in all groups, the squat-shaped were the lowest. (4) The big toes were longer than the second toes in most people of 6 groups,but without significant sexual differences in each groups. (5) There were no significant correlations between 4 characters.

The data of 72 families were analyzed by the method of proband's sib and the method of segregation analysis. The results showed that the hereditary mode of rolling tongue or pointed tongue is the dominant heredity of single gene of autosome, and the positive type of them is the dominant character. Twisting tongue is the recessive heredity of single gene of autosome,while the positive type is the recessive character. Present study suggested, although environmental might affect these characters,hereditary factors seemed to be dominant.

The gene frequency of 12 characters was reported in Ewenki, Oroqen, Daur, Mongol and Han nationalities in Inner Mongolia, and compared among these nationalities. The result indicated that the difference of Mongoloid fold among nationalities was significant, followed by the Folding tongue, while the difference of Handedness and Nasal profile was relatively insignificant.

We report the cloning and characterization of a gene called cotF from Bacillus subtilis that encodes alkali-soluble polypeptides of 5 and 8 kDa that are components of the spore coat. The 5- and 8-kDa polypeptides are generated by proteolytic cleavage of the primary product of the cotF gene, which is 160 codons in length and is capable of encoding a polypeptide of 19 kDa. Amino acid sequence analysis indicates that the 5-kDa species is derived from the NH2-terminal portion of the primary gene product and that the 8-kDa species is derived from the COOH-terminal portion. A mutant bearing an in vitro-constructed cotF null mutation produced normal-looking spores that contained an apparently complete set of coat proteins except for the absence of the 5- and 8-kDa polypeptides. The map position of cotF is 349 degrees. Transcription of cotF commenced coincidently (during h 6 of sporulation) with genes known to be under the control of sporulation transcription factor sigma kappa.

Endospores of the Gram-positive bacterium Bacillus subtilis are encased in a tough protein shell known as the coat. The coat is composed of a dozen or more different structural proteins. We report the identification of and studies on the regulation of promoters governing the expression of coat protein (cot) genes designated B to E encoding polypeptides of 59, 12, 11 and 24 kDa, respectively. We show that transcription of genes B, C and D is governed by single promoters and that transcription of gene E is governed by tandem promoters designated P1 and P2. In extension of recent work on the transcription of cot gene A and the mother-cell regulatory genes gerE, sigK and spoIIID, we show that genes involved in coat formation are turned on in a regulatory cascade of at least four co-ordinately controlled gene sets. The cascade consists of: cotE as transcribed from its P1 promoter and spoIIID, which are turned on during hours three to four of sporulation; cotE as transcribed from its P2 promoter and sigK, which are turned on during hour five by the appearance of the product (a small DNA-binding protein) of spoIIID; cotA, cotD and gerE, which are turned on during hours five to six by the appearance of the product (sigma factor sigma K) of sigK; and cotB and cotC, which are turned on during hour seven by the appearance of the product (an inferred DNA-binding protein) of gerE. The cascade is hierarchical in that the first three gene sets each contain the regulatory gene that turns on the expression of the next gene set in the pathway. We also show that the level of expression of a member (cotC) of the terminal class of gene expression is strongly influenced by medium and that this effect directly or indirectly depends on the product of sporulation gene spoIV A.

Endospores of the Gram-positive bacterium Bacillus subtilis are encased in a tough protein shell, known as the coat, that consists of a dozen or more different polypeptides. We have cloned structural genes designated cotA, cotB, cotC and cotD that encode spore coat proteins of Mr 65,000, 59,000, 12,000 and 11,000, respectively. These genes were cloned by using as hybridization probes synthetic oligonucleotides that were designed on the basis of partial NH2-terminal sequence determinations of the purified coat proteins. To determine the location of the cot genes on the chromosome and to study their function genetically, we tagged each gene by insertion of a chloramphenicol-resistance determinant (cat) within its coding sequence. We then replaced each wild-type cot gene in the chromosome with the corresponding, insertionally inactivated gene. Genetic mapping experiments showed that cotA, cotB, cotC and cotD were located at 52 degrees, 290 degrees, 168 degrees and 200 degrees, respectively, on the B. subtilis chromosome. None of the cot::cat insertion mutants were Spo-, but spores of the cotD mutant were found to germinate somewhat more slowly than did wild-type spores, and the cotA mutant was found to be blocked in the appearance of the brown pigment characteristic of colonies of wild-type sporulating cells. Physical and genetic experiments established that cotA was identical to a previously identified gene called pig, known to be responsible for sporulation-associated pigment production. Spores from all four insertion mutants exhibited the wild-type pattern of coat polypeptides, except for the absence in each instance of the corresponding product of the cot gene that had been insertionally inactivated.

Endospores of Bacillus subtilis are encased in a two-layer protein shell known as the coat, which consists of a lammellar-like inner layer and an electron-dense outer layer. We report the cloning of the structural gene (designated cotE) for an alkali-soluble coat protein of 24 kD and show that the cotE gene product is a morphogenic protein required in the assembly of the outer coat. The nucleotide sequence of cotE reveals an open reading frame capable of encoding a 181-residue-long polypeptide of 21 kD. A cotE mutant was created by replacing the chromosomal gene, which was located at 145 degrees on the chromosome, with an in vitro constructed, deletion-mutated gene. The resulting cotE mutant formed normal-looking (optically refractile) spores that were heat resistant but were sensitive to lysozyme and somewhat impaired in germination. Ultrastructural analysis indicated that the mutant spores lacked the electron-dense outer layer of the coat but retained a normal-looking inner coat. The mutant spores were pleiotropically deficient in several coat proteins, including the product of cotE and the products of previously cloned cot genes A-C. Based on experiments in which expression of the cotA and cotC genes was found to be unimpaired in cotE mutant cells, we infer that the cotE gene product is involved in the assembly of the products of cotA-cotC, and certain other proteins into the electron-dense outer layer of the coat.