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Antrodia cinnamomea (Ac), also known as "Niu-Chang-Chih" in Chinese, is a valuable fungus that has been widely used as medicine and food among indigenous people in Taiwan. Ac is rich in polysaccharides (Ac-PS), making it a promising candidate for adjunctive therapy in cancer and inflammation conditions. There are two types of Ac-PS: general (non-sulfated) PS (Ac-GPS) and sulfated PS (Ac-SPS). This review highlights that both Ac-GPS and Ac-SPS possess immunomodulatory, anti-inflammatory and anti-cancer properties. Each type influences interleukin signaling pathways to exert its anti-inflammatory effects. Ac-GPS is particularly effective in alleviating inflammation in the brain and liver, while Ac-SPS shows its efficacy in macrophage models. Both Ac-GSP and Ac-SPS have demonstrated anti-cancer effects supported by in vitro and in vivo studies, primarily through inducing apoptosis in various cancer cell lines. They may also synergize with chemotherapy and exhibit anti-angiogenic properties. Notably, Ac-SPS appears to have superior anti-cancer efficacy, potentially due to its sulfate groups. Furthermore, Ac-SPS has been more extensively studied in terms of its mechanisms and effects on lung cancer compared with Ac-GPS, highlighting its significance in cancer research. In addition, Ac-SPS is often reported for its ability to activate macrophage-mediated responses. Clinically, Ac-GPS has been used as an adjunctive therapy for advanced lung cancer, as noted in recent reports. However, given the numerous studies emphasizing its anti-cancer mechanisms, Ac-SPS may exhibit greater efficacy, warranting further investigation. This review concludes that Ac-derived Ac-GPS or Ac-SPS have the potential to be developed into functional health supplements or adjunctive therapies, providing dual benefits of anti-inflammatory and anti-cancer effects., Competing Interests: Conflicts of interest: The authors declare that they have no conflicts of interest related to the subject matter or materials discussed in this article., (Copyright © 2024, the Chinese Medical Association.)

Objective: Two classes of spiro[4 H -pyran-3,3'-oxindole] derivatives were prepared via the one pot reaction of chain diketones (1-phenyl-1,3-butanedione or dibenzoyl methane), substituted isatins and malononitrile successfully catalyzed by a Tröger's base derivative 1b (5,12-dimethyl-3,10-diphenyl-bis-1 H -pyrazol [b,f ][4,5]-1,5-diazadicyclo[3.3.1]-2,6-octadiene). The antibacterial activity of products against three wild-type bacteria ( B. subtilis, S. aureus , and E. coli) and two resistant strains (Methicillin-resistant S. aureus (18H 8 ) and E. coli carrying the BlaNDM-1 gene (18H 5 )) was evaluated using the minimum inhibitory concentration (MIC).., Methods: 1-Phenyl-1,3-butanedione 2 or dibenzoylmethane 2' (0.42 mmol), substituted isatin 3 (0.4 mmol), malononitrile 4 (0.8 mmol), Tröger's base derivative 1b (0.08 mmol), and 10 mL of acetonitrile were added to a 50 mL round bottom flask and refluxed. After the completion (TLC monitoring), water (10 mL) was added to the reaction mixture; pH = 7 was adjusted with saturated NaHCO 3 (aq.), and the mixture was extracted with CH 2 Cl 2 (50 mL × 3). Organic layers were combined and dried with anhydrous Na 2 SO 4 ; the solvent was removed under vacuum, and the residue was purified by column chromatography (V DCM : V MeOH = 80: 1) to afford product 5. The antibacterial activity was tested by the MTT method., Results: Seventeen spiro[4 H -pyran-3,3'-oxindole] derivatives were synthesized through the reaction of chain diketones (1-phenyl-1,3-butanedione or dibenzoyl methane), substituted isatins, and malononitrile in one-pot in medium to high yields. Four compounds showed antibacterial activity, and two of them showed the same activity as the positive control Ceftazidime on S. aureus (MIC = 12.5 μg/mL)., Conclusion: Two classes of spiro[4 H -pyran-3,3'-oxindole] derivatives were prepared, and their antibacterial activity was evaluated. Tröger's base derivative 1b (5,12-dimethyl-3,10-diphenyl-bis-1 H -pyrazol[ b,f ][4,5]- 1,5-diazadicyclo[3,3,1]-2,6-octadiene) was used as an efficient organocatalyst for the reaction of low reactive chain diketones (1-phenyl-1,3-butanedione or dibenzoyl methane), substituted isatins, and malononitrile in one-pot successfully and effectively by providing multiple active sites and alkaline environment. By the theoretical calculation, we explained the possible reaction sequence and mechanism. Due to the superiority and high efficiency of the TB framework as an organocatalyst, the reaction showed many advantages, including mild reaction conditions, low catalyst loading, and a wide substrate range. It expanded the application of Tröger's base to the multicomponent reaction in organocatalysis. Some products were screened due to their high antibacterial activity in vitro , showing their potential in new antibacterial drug development., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Objective: To explore the possible mechanism of fluid shear stress on human nucleus pulposus cells based on label-free proteomics technology., Methods: The human nucleus pulposus cell line was purchased and subcultured in vitro. The Flexcell STR-4000 multiflow field cell fluid shear stress loading culture system was used to apply continuous laminar fluid shear stress (12 dyne/cm 2 , 45 mins) to the monolayer adherent cells. Those without mechanical loading were used as the control group, and those subjected to fluid shear loading were used as the experimental group. Differential protein expression was identified using mass spectrometry identification technology, and bioinformatics analysis was performed using Gene Ontology GO (Gene Ontology) and Kyoto Encyclopedia of Genes and Genomes KEGG (Kyoto Encyclopedia of Genes and Genomes)., Results: The proteomics results of the experimental group and the control group showed that the total number of mass spectra was 638653, the number of matched mass spectra was 170110, the total number of identified peptides was 32050, the specific peptide was 30564, and the total number of identified proteins was 4745. Comparing the two groups, 47 proteins were significantly differentially expressed, namely, 25 upregulated proteins and 22 downregulated proteins. Bioinformatics analysis showed that significantly different proteins were mainly manifested in cellular process, biological regulation, metabolic process, binding, catalytic activity, cellular components (cell part), organelle part (organelle part), and other molecular biological functions., Conclusion: Using proteomics technology to screen human nucleus pulposus cells after fluid shear stress loading, the differential protein expression provides a basis for further exploration of the mechanism of mechanical factors on nucleus pulposus., Competing Interests: All the authors declare no conflict of interests., (Copyright © 2022 Liang-yu Xie et al.)

Objective: Angiopoietin-like protein 4 (ANGPTL4), encoding a glycosylated secreted protein, has been reported to be closely related to many kinds of diseases, including diabetes, tumor, and some musculoskeletal pathologies, such as rheumatoid arthritis, osteoarthritis, and osteoporosis. The aim of the current study is to investigate the role of ANGPTL4 in intervertebral disc degeneration and analyze the association of ANGPTL4 expression with Pfirrmann grades., Methods: A total of 162 nucleus pulposus tissues were collected from lumbar intervertebral disc herniation patients undergoing interforaminal endoscopic surgery. Real-time quantitative PCR and western blot were performed to determine the mRNA and protein expression of ANGPTL4 in nucleus pulposus samples. Statistical analysis was performed to analyze the association of ANGPTL4 expression with Pfirrmann grades., Results: Based on the clinical data of 162 patients, results showed that Pfirrmann grades were significantly associated with patients' age ( r = 0.162, P = 0.047) and were not significantly associated with patients' gender ( P > 0.05). RT-qPCR and western blot results showed that the mRNA ( r = 0.287, P < 0.05) and protein ( r = 0.356, P < 0.05) expressions of ANGPTL4 were both closely associated with Pfirrmann grades. The expression of ANGPTL4 was remarkably increased in the groups of high IVDD Pfirrmann grades., Conclusion: The results demonstrated that ANGPTL4 expression was positively associated with the Pfirrmann grades and the severity of intervertebral disc degeneration. ANGPTL4 may be served as a candidate biomarker for intervertebral disc degeneration., Competing Interests: All the authors declare no conflict of interests., (Copyright © 2021 Fan-jie Liu et al.)

Background: Colla Cornus Cervi (CCC) has been used as a traditional Chinese medicine in the treatment of osteoporosis and osteonecrosis of the femoral head. However, the bioavailability of CCC is seriously limited owing to its large molecular weight and complex ingredients. In the present study, antler polypeptide was separated from CCC, and the effects of antler polypeptide on rat bone marrow mesenchymal stem cells (BMSCs) were investigated., Methods: Antler polypeptide was separated from Colla Cornus Cervi by ultrafiltration into different samples according to the molecular weight. The total peptide content of these samples was determined by the biuret method. The content of antler polypeptide in different samples was quantified by high-performance liquid chromatography (HPLC). The effects of antler polypeptide at different concentrations on the proliferation, cell cycle, alkaline phosphatase activity, and BMP7 expression of BMSCs were investigated., Results: Antler polypeptide was separated by ultrafiltration into different samples: A (molecular weight <800 Da), B (molecular weight 800-1500 Da), and C (molecular weight >1500 Da). The total peptide contents of A, B, and C were 0.602 mg/mL, 8.976 mg/mL, and 38.88 mg/mL. Antler polypeptide B eluted at 14.279∼15.351 min showed that the content of antler polypeptide was significantly higher than that of A and C with a peak area of 933.80927. The BMSCs proliferation rate (84.66%) of polypeptide B was the highest at the concentration of 1.578 × 10 -2  g/mL. Antler polypeptide B significantly promoted the proliferation of BMSCs with a proliferation index of 38.68%, which was significantly higher than that of the other groups. Antler polypeptide B significantly enhanced the activity of alkaline phosphatase in BMSCs compared to that of the blank group ( P < 0.001). Antler polypeptide B increased the BMP7 protein expression in BMSCs., Conclusions: Results suggested that antler polypeptide may promote the proliferation and osteogenic differentiation of BMSCs. Our study lays an experimental foundation for the further development and application of antler polypeptide in medicine., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Ping Wang et al.)

Objective: To explore the biomechanical characteristics of "three-dimensional balanced manipulation" for the treatment of cervical spondylotic radiculopathy(CSR)., Methods: A CSR patient was treated with "three-dimensional balanced manipulation", and the mechanical changes during the manipulation were monitored by mechanical testing system. Using spiral CT to scan the neck of the patient to obtain DICOM data. The three-dimensional finite element model of cervical spondylotic radiculopathy was established by using Mimics software, Geographic Studio software. The "three-dimensional balance manipulation" was simulated and loaded, and the mechanical parameters of each part were replaced into the finite element model, and the finite element analysis was carried out by using ANSYS software to study the internal stress changes and displacement deformation of vertebral body and intervertebral disc under the action of "three-dimensional balance manipulation"., Results: The established C 3 -C 7 finite element model of the CSR patient consisted of 5 vertebrae, 4 intervertebral discs and 3 ligaments, involving 153 471 nodes and 64 978 units. The stress of C 3 -C 7 vertebral body was mainly located in anterior and root of C 5 spinous processes, arch, vertebral arch and the combination of the two after full loading of manipulation, and the maximum stress was 17.781 MPa. The deformation sites were mainly concentrated in articular processes and anterior transverse processes of C 3 , superior articular processes and transverse processes of C 4 , articular processes of C 5 . The stress of C 3 -C 7 intervertebral disc mainly distributed in the anterior part of C 3, 4 intervertebral disc and the nucleus pulposus of C 4, 5 and C 5, 6 . The displace mentextended to the middle and posterior part of C 3, 4 nucleus pulposus, around the nucleus of C 4, 5 and C 5, 6 and anterior part of cervical intervertebral disc., Conclusion: The establishment of three-dimensional finite element model of C 3 -C 7 cervical spondylotic radiculopathy can simulate the geometry and material properties of cervical spine, and also accurately reflects the biomechanical characteristics of cervical spine, verifys the internal mechanism of "three-dimensional balanced manipulation" on CSR, proves the safety and effectiveness of treatment, guides more standardized manipulation, and avoids medical accidents.

Simultaneous transcriptome analyses of both host plants and pathogens, and functional validation of the identified differentially expressed genes (DEGs) allow us to better understand the mechanisms underlying their interactions. Here, we analyse the mixed transcriptome derived from Botrytis cinerea (the causal agent of grey mould) infected tomato leaves at 24 hr after inoculation, a critical time point at which the pathogen has penetrated and developed in the leaf epidermis, whereas necrotic symptoms have not yet appeared. Our analyses identified a complex network of genes involved in the tomato-B. cinerea interaction. The expression of fungal transcripts encoding candidate effectors, enzymes for secondary metabolite biosynthesis, hormone and reactive oxygen species (ROS) production, and autophagy-related proteins was up-regulated, suggesting that these genes may be involved in the initial infection processes. Specifically, tomato genes involved in phytoalexin production, stress responses, ATP-binding cassette transporters, pathogenesis-related proteins, and WRKY DNA-binding transcription factors were up-regulated. We functionally investigated several B. cinerea DEGs via gene replacement and pathogenicity assays, and demonstrated that BcCGF1 was a novel virulence-associated factor that mediates fungal development and virulence via regulation of conidial germination, conidiation, infection structure formation, host penetration, and stress adaptation. The fungal infection-related development was controlled by BcCGF-mediated ROS production and exogenous cAMP restored the mutant infection-related development. Our findings provide new insights into the elucidation of the simultaneous tactics of pathogen attack and host defence. Our systematic elucidation of BcCGF1 in mediating fungal pathogenesis may open up new targets for fungal disease control., (© 2020 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Objective: The aim was to investigate the role of microRNA-26b-5p in regulating mesenchymal stem cells (MSCs) differentiation to type II of alveolar epithelial cells (AECII) in the disease course of neonatal respiratory distress syndrome (NRDS)., Materials and Methods: MSCs were first derived from rat bone marrow. In vitro induction of MSCs differentiation to AECII was conducted by SAGM. The mRNA levels of microRNA-26b-5p, Wnt5a, and AECII-related genes (Occludin, KGF, CK18, SpA, SpB, and SpC) during the process of cell differentiation were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Enzyme-linked immunosorbent assay (ELISA) was conducted for detecting levels of inflammatory factors tumor necrosis factor-α (TNF-α), interferon-α (INF-α), and interleukin-1 (IL-1) in cell supernatant. Dual-luciferase reporter gene assay was then carried out to verify the regulatory effect of microRNA-26b-5p on Wnt5a. MicroRNA-26b-5p expression in serum samples of NRDS neonates and healthy neonates was detected by qRT-PCR as well., Results: MicroRNA-26b-5p was overexpressed in NRDS neonates than those of healthy neonates. Besides, microRNA-26b-5p was highly expressed in the process of MSCs differentiation to AECII. MicroRNA-26b-5p overexpression remarkably inhibited AECII differentiation and Wnt5a expression. Levels of TNF-α, INF-α, and IL-1 in cell supernatant during differentiation induction were elevated. The regulatory effects of microRNA-26b-5p on AECII differentiation, Wnt5a expression, and inflammatory response were reversed by Wnt5a overexpression., Conclusions: MicroRNA-26b-5p inhibits MSCs differentiation to AECII via inhibiting Wnt5a expression through the Wnt pathway.

The process of initiation of host invasion and survival of some foliar phytopathogenic fungi in the absence of external nutrients on host leaf surfaces remains obscure. Here, we demonstrate that gluconeogenesis plays an important role in the process and nutrient-starvation adaptation before the pathogen host invasion. Deletion of phosphoenolpyruvate carboxykinase gene BcPCK1 in gluconeogenesis in Botrytis cinerea, the causative agent of grey mould, resulted in the failure of the ΔBcpck1 mutant conidia to germinate on hard and hydrophobic surface and penetrate host cells in the absence of glucose, reduction in conidiation and slow conidium germination in a nutrient-rich medium. The wild-type and ΔBcpck1 conidia germinate similarly in the presence of glucose (higher concentration) as the sole carbon source. Conidial glucose-content should reach a threshold level to initiate germination and host penetration. Infection structure formation by the mutants displayed a glucose-dependent fashion, which corresponded to the mutant virulence reduction. Exogenous glucose or complementation of BcPCK1 completely rescued all the developmental and virulence defects of the mutants. Our findings demonstrate that BcPCK1 plays a crucial role in B. cinerea pathogenic growth and virulence, and provide new insights into gluconeogenesis mediating pathogenesis of plant fungal pathogens via initiation of conidial germination and host penetration., (© 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Botrytis cinerea is a necrotrophic plant fungal pathogen that annually causes enormous economic losses worldwide. The ribosome is an organelle for cellular protein biosynthesis. However, little is known about how the ribosome operates as a machine to mediate microbial pathogenesis. Here, we demonstrate that Nop53, a late-acting factor for 60S ribosomal subunit maturation, is crucial for the pathogen's development and virulence. BcNop53 is functionally equivalent to yeast nop53p. Complementation of BcNOP53 completely restored the growth defect of the yeast Δnop53 mutant. BcNop53 is located in nuclei and disruption of BcNOP53 also dramatically impaired pathogen growth. Deletion of BcNOP53 blocked infection structure formation and abolished virulence of the pathogen, possibly due to reduced production of reactive oxygen species. Moreover, loss of BcNOP53 impaired pathogen conidiation and stress adaptation, altered conidial and sclerotial morphology, retarded conidium and sclerotium germination as well as reduced the activities of cell-wall degradation-associated enzymes. Sclerotium production was, however, increased. Complementation with the wild-type BcNOP53 allele rescued defects found in the ΔBcnop53 mutant. Our work establishes a systematic elucidation of Nop53 in regulating microbial development and pathogenesis, provides novel insights into ribosomal processes that regulate fungal pathogenesis, and may open up new targets for addressing fungal diseases., (© 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Neuroinflammation triggered by activation of glial cells plays an important role in the pathophysiology of several neurodegenerative diseases including Parkinson's disease (PD). Besides microglia, astrocytes are also critical in initiating and perpetuating inflammatory process associated with PD. Heat shock protein 70 (Hsp70) is originally described as intracellular chaperone, however, recent study revealed that it had anti-inflammatory effects as well. The present study is designed to investigate whether Hsp70 mediates neuroinflammation in astrocytes. By employing α-synuclein (α-Syn) (A53T) aggregates on primary cultured astrocytes of rats, we found that astrocytes were activated and neuroinflammatory response was triggered, as indicated by over-expression of glial fibrillary acidic protein (GFAP), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), increased production of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β). The data also showed that the neuroinflammatory response accompanied up-regulated Hsp70 expression. Moreover, over-expression of Hsp70 through transfection of Hsp70 cDNA plasmids could significantly reduce the production of TNF-α, IL-1β, and the expression of GFAP, COX-2 as well as iNOS. While inhibition of Hsp70 by VER155008 exacerbated neuroinflammatory response in astrocytes challenged by α-Syn aggregates. Further mechanistic study indicated that c-Jun N-terminal kinase (JNK) and nuclear factor-κB (NF-κB) signalings were responsible for the neuroinflammation, which was also regulated by Hsp70. These findings demonstrated that Hsp70 was an important modulator in astrocytes induced inflammation, and up-regulation of Hsp70 might be a potential regulating approach for neuroinflammation-related neurodegenerative diseases, such as PD., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Anaplasmosis and theileriosis are significant tick-borne diseases threatening the livestock industry worldwide. In the present study, we screened 127 cattle and 115 sheep blood DNA samples from northeastern China for Theileria and Anaplasma pathogens by polymerase chain reaction (PCR) using species-specific primers. The result showed that only Theileria orientalis and Anaplasma ovis were detected, with a prevalence of 2.9% for T. orientalis in cattle and 57.4% for A. ovis in sheep. Fragments of Anaplasma ovis major surface protein 4 (AoMSP4) and Theileria orientalis major piroplasm surface protein (ToMPSP) genes were sequenced for phylogenetic analysis. Sequence analysis showed that the AoMSP4 gene was conserved, with 100% sequence identity value among sheep samples. However, the ToMPSP gene was relatively diverse, with sequence identity ranging from 87.6%-99l.0% among cattle samples. Phylogenetic analysis showed that the ToMPSP gene sequences isolated from 4 cattle samples were classified into type 1, type 2 and type 7, while the AoMSP4 gene sequences obtained from 66 sheep were classified into genotype I, according to the neighbour-joining distance method. This study provides important data for understanding the epidemiology of tick-borne diseases and genetic diversity of these pathogens in the northeast region of China.

Bovine theileriosis is a tick-borne disease that is hampering the development of the domestic cattle industry in northern China. This study involved a molecular survey of bovine Theileria species in 137 blood samples from cattle in the Jilin province of China. The DNA samples were screened by species-specific 18S rRNA PCR. Results revealed that 19.7% (27/137), 17.5% (24/137) and 10.9% (15/137) were found to be infected with Theileria sinensis, Theileria orientalis, respectively. Mixed infection was found in 8.8% (12/137). The overall detection rates of Baishan, Yanji, Jilin and Liaoyuan districts was 60.0%, 17.5%, 5.3% and 0%, respectively. There is little information on the detection and distribution of bovine Theileria species in northern China. Therefore, this study provides important data for understanding the epidemiology of Theileria species and designing appropriate approaches for the diagnosis and control of bovine theileriosis in northern China.

Objective To investigate the mechanism of non-receptor tyrosine kinase Src regulating neuroinflammation through phosphatase and tensin homology protein(PTEN)in microglia. Methods BV2 cells were incubated with PTEN inhibitor bpv(HOpic)for 2 hours,and then added with lipopolysaccharide(LPS)to induce neuroinflammation,Western blot was performed to determine the expression of phosphorylated protein kinase B(Akt)to investigate the activity of PTEN. Enzyme-linked immunosorben assay(ELISA)was used to determine the release of tumor necrosis factor α(TNF-α)to assess neuroinflammation.After PTEN inhibitor or Src specific small interfering RNA was added,the change of neuroinflammation was evaluated to study the mechanism of Src regulating neuroinflammation. Results LPS induced significant neuroinflammation in BV2 cells,as indicated by significantly increased expression of p-Akt and release of TNF-α(P<0.001).The PTEN inhibitor signficantly increased Akt phosphorylation(P<0.05)and TNF-α release(P<0.001)in LPS-induced BV2 cells compared to simply LPS-induced cells.The Src small interfering RNA significantly decreased the release of TNF-α(P<0.001)and inhibited PTEN(P<0.001)and Akt(P<0.001)phosphorylation. Conclusion Src kinase may regulate neuroinflammtion response in BV2 cells by regulating the phosphorylation of PTEN.

Numerous drug treatments are available for Parkinson's disease (PD), an age-related neurodegenerative disease, but most cause serious side effects. Therefore, novel therapeutic strategies that halt disease progression and allow for long-term administration are urgently needed. Neuroinflammation critically contributes to the pathogenesis of PD. Here, we report the discovery and optimization of phloroglucinol derivatives, a novel class of anti-neuroinflammatory compounds. Structural modifications of the hit compound 3-methyl-1-(2,4,6-trihydroxyphenyl)butan-1-one produced 43 derivatives, including a preclinical candidate (compound 21), that exhibited potent in vitro anti-neuroinflammatory effects, good blood-brain barrier penetration, and desirable safety margins in mice at a median lethal dose (LD 50 ) >5000 mg/kg. Its in vivo efficacy was demonstrated in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)- and MPTP/probenecid (prob)-induced subacute and chronic PD models, respectively, and α-synuclein transgenic mice. Mechanistic studies revealed neuroinflammation inhibition by targeting Src/phosphatase and tensin homologue deleted on chromosome 10 (PTEN)/Akt signaling might be promising. We highlighted the potential usefulness of phloroglucinol derivatives in PD treatment.

Microglia are the resident innate immune cells in the brain. Under endogenous or exogenous stimulates, they become activated and play an important role in the neurodegenerative diseases. Microglial phagocytosis is a process of receptor-mediated engulfment and degradation of apoptotic cells. In addition, microglia can phagocyte brain-specific cargo, such as myelin debris and abnormal protein aggregation. However, recent studies have shown that microglia can also phagocyte stressed-but-viable neurons, causing loss of neurons in the brain. Thus, whether microglial phagocytosis is beneficial or not in neurodegenerative disease remains controversial. This article reviews microglial phagocytosis related mechanisms and its potential roles in neurodegenerative diseases, with an attempt to provide new insights in the treatment of neurodegenerative diseases.

Purpose: Photoreceptor neuronal degenerations are common, incurable causes of human blindness affecting 1 in 2000 patients worldwide. Only half of all patients are associated with known mutations in over 250 disease genes, prompting our research program to identify the remaining new genes. Most retinal degenerations are restricted to the retina, but photoreceptor degenerations can also be found in a wide variety of systemic diseases. We identified an X-linked family from Sri Lanka with a severe choroidal degeneration and postulated a new disease entity. Because of phenotypic overlaps with Bietti's crystalline dystrophy, which was recently found to have systemic features, we hypothesized that a systemic disease may be present in this new disease as well., Methods: For phenotyping, we performed detailed eye exams with in vivo retinal imaging by optical coherence tomography. For genotyping, we performed whole exome sequencing, followed by Sanger sequencing confirmations and cosegregation. Systemic investigations included electron microscopy studies of peripheral blood cells in patients and in normal controls and detailed fatty acid profiles (both plasma and red blood cell [RBC] membranes). Fatty acid levels were compared to normal controls, and only values two standard deviations above or below normal controls were further evaluated., Results: The family segregated a REP1 mutation, suggesting choroideremia (CHM). We then found crystals in peripheral blood lymphocytes and discovered significant plasma fatty acid abnormalities and RBC membrane abnormalities (i.e., elevated plasmalogens). To replicate our discoveries, we expanded the cohort to nine CHM patients, genotyped them for REP1 mutations, and found the same abnormalities (crystals and fatty acid abnormalities) in all patients., Conclusions: Previously, CHM was thought to be restricted to the retina. We show, to our knowledge for the first time, that CHM is a systemic condition with prominent crystals in lymphocytes and significant fatty acid abnormalities.

In the Tris-HCl buffer solution with pH was 7.40, the interaction between glipizide (Gli) and bovine serum albumin (BSA) was investigated by classical fluorescence spectroscopy with the change of protein as investigation object and elastic scattering fluorescence spectrometry with the change of drugs as investigation object at 293 K and 303 K, the conclusions of the two methods were consistent. Results showed that Gli could quench the intrinsic fluorescence of BSA, and the quenching mechanism was a dynamic quenching process. The hydrophobic force played an important role in the conjugation reaction between BSA and Gli, the binding site mainly located in BSA hydrophobic region and the number of binding site (n) in the binary system was approximately to 1. The values of Hill's coefficients were less than 1, which indicated the weak negative cooperativity in BSA-Gli system. The binding constant (Ka) obtained by elastic scattering fluorescence spectrometric was much larger than the one obtained by classical fluorescence spectroscopy, indiciating that it was more accurate and reasonable when using the change of drug's fluorescence as the research object. At last, the scientificalness of the new method based on elastic scattering fluorescence spectrometric was verified by ultraviolet spectroscopy. The research results showed that there existed insufficiency in analysis of the interaction of drug with protein by classical fluorescence spectroscopy with the change of protein as investigation object, and the fluorescence spectrogram only reflected partial information of the interaction between drug and protein, while the interaction between drug and protein could be better expressed by elastic scattering fluorescence spectrometry with the change of drugs as investigation object.

Aim: To prepare functional monoclonal antibodies(mAb)against recombinant human Flt-1(rhFlt-1)., Methods: A cell line stable secreting mAb was established by using FLT-1 extracellular domain III as antigen and hybridoma technique. Then it was purified in large-scale from mouse ascites by protein G affinity chromatography. The characteristics of mAb were then determined by ELISA, Western blotting and FACS., Results: The immunoglobin subtype of mAb XA12 was IgG1 with kappa (κ) light chains, and it could recognize rhFlt-1 specifically. Furthermore, mAb XA12 could bind to rhFlt-1with high affinity (K=1.28±0.05 nmol/L). It could also be used to detect Flt-1-positive cells, such as human umbilical vascular endothelial cells (HUVECs) and K562/A02 in a dose-dependent fashion., Conclusion: A hybridoma cell line secreting functional anti-rhFlt-1 mAb was successfully prepared. The antibody can be used to study the function of Flt-1 and further potentially optimized for clinical purpose.

Objective: The histopathologic abnormality underlying ascending aortic aneurysm and dissection is medial degeneration, a lesion that is described as the noninflammatory loss of smooth muscle cells and elastic fibers. This study sought to determine whether inflammatory cells are present in medial degeneration and assess any possible contribution of these cells to apoptosis of smooth muscle cells., Methods: Aortic specimens were obtained from patients undergoing prophylactic surgical repair of an ascending aortic aneurysm (n = 9) and type A dissection (n = 7), along with control patients dying of causes unrelated to aortic disease (n = 5). Immunohistochemical staining was performed to evaluate the presence of lymphocytes and macrophages, and markers of apoptosis were assessed in the aortas of patients with ascending aortic aneurysm and dissection., Results: Immunohistochemical study indicated significantly more CD3+ cells in the aortas of patients with aneurysms or dissections than in control aortas (P = .020 and P = .0022, respectively). In addition, aortas of patients with aneurysms or dissections had more CD68+ cells (P = .01 and P = .005, respectively). CD3+ cells were localized in the media and surrounding the vasa vasorum in the adventitia. Cells yielding a positive result on in situ terminal transferase-mediated deoxyuridine triphosphate nick end-labeling were found in increased numbers in the aortas of patients with aneurysms or dissections relative to control aortas (P = .005 and P = .002, respectively). Furthermore, Fas and FasL were increased in the aortic samples from patients with aneurysms and dissections relative to control aortas., Conclusion: The coexistence of inflammatory cells with markers of apoptotic vascular cell death in the media of ascending aortas with aneurysms and type A dissections raises the possibility that activated T cells and macrophages may contribute to the elimination of smooth muscle cells and degradation of the matrix associated with thoracic aortic aneurysms and dissections.

Fibrillin-1 is synthesized as a proprotein that undergoes proteolytic processing in the unique C-terminal domain by a member of the PACE/furin family of endoproteases. This family of endoproteases is active in the trans-Golgi network (TGN), but metabolic labeling studies have been controversial as to whether profibrillin-1 is processed intracellularly or after secretion. This report provides evidence that profibrillin-1 processing is not an intracellular event. Bafilomycin A(1) and incubation of dermal fibroblasts at 22 degrees C were used to block secretion in the TGN to confirm that profibrillin-1 processing did not occur in this compartment. Profibrillin-1 immunoprecipitation studies revealed that two endoplasmic reticulum-resident molecular chaperones, BiP and GRP94, interacted with profibrillin-1. To determine the proprotein convertase responsible for processing profibrillin-1, a specific inhibitor of furin, alpha-1-antitrypsin, Portland variant, was both expressed in the cells and added to cells exogenously. In both cases, the inhibitor blocked the processing of profibrillin-1, providing evidence that furin is the enzyme responsible for profibrillin-1 processing. These studies delineate the secretion and proteolytic processing of profibrillin-1, and identify the proteins that interact with profibrillin-1 in the secretory pathway., (Copyright 2003 Wiley-Liss, Inc.)

The purpose of this study was to determine whether mutations in the Bax gene play a role in the development of ovarian endometrioid carcinoma with a microsatellite instability phenotype. We analyzed a total of 60 tumor specimens, 49 ovarian endometrioid carcinomas and 11 concurrent endometrial endometrioid carcinomas from 49 patients. Fourteen ovarian endometrioid carcinomas and 6 endometrial endometrioid carcinomas showed a microsatellite instability-high phenotype. Tumor and normal-tissue specimens from eight patients with a microsatellite instability-high phenotype colorectal carcinoma were included in this study as controls. The presence or absence of a mutation in the poly (G) 8 tract of the Bax gene was determined by polymerase chain reaction followed by direct DNA sequence analysis. A 1-base pair deletion at the poly (G) 8 tract and no expression of Bax and Bcl-2 proteins were identified in one microsatellite instability-high endometrial endometrioid carcinoma. Immunohistochemical staining for Bax and Bcl-2 proteins was negative on the tumor specimen that had this 1-base pair deletion. No mutations were found in the synchronous microsatellite instability-high ovarian endometrioid carcinoma from the same patient. In contrast, four (50%) of the eight microsatellite instability-high sporadic colorectal carcinomas had a mutation in the poly (G) 8 tract. Although Bax plays an important role in carcinogenesis of the colorectum with microsatellite instability-high phenotype, Bax may not play a direct role in the genesis of ovarian endometrioid carcinoma, regardless of microsatellite instability status.

We describe a 14-year-old male with dissection of the descending aorta, bilateral iris hypoplasia, striae distensae and brachytelephalangy, the latter being most marked in the thumbs. Inguinal herniae and a patent ductus arteriosus were surgically repaired in infancy. The pattern of abnormalities may constitute a previously undescribed syndrome. The proband died suddenly at the age of 17 years.

Dermal fibroblasts from a 13-yr-old boy with isolated skeletal features of the Marfan syndrome were used to study fibrillin synthesis and processing. Only one half of the secreted profibrillin was proteolytically processed to fibrillin outside the cell and deposited into the extracellular matrix. Electron microscopic examination of rotary shadowed microfibrils made by the proband's fibroblasts were indistinguishable from control cells. Sequencing of the FBN1 gene revealed a heterozygous C to T transition at nucleotide 8176 resulting in the substitution of a tryptophan for an arginine (R2726W), at a site immediately adjacent to a consensus sequence recognized by a cellular protease. Six other individuals in the proband's family had the FBN1 mutation that segregated with tall stature. None of the affected individuals have cardiac or ocular manifestations of the Marfan syndrome. This mutation identifies a putative site for profibrillin to fibrillin processing, and is associated with isolated skeletal features of the Marfan syndrome, indicating that the FBN1 gene is one of the genes that determines height in the general population. The cellular effect of the mutation may be equivalent to a "null" FBN1 allele and may define the phenotype associated with FBN1 "null" alleles.

Elevated serum ELISA IgG antibodies to rubella virus (RV) were found by three independent determinations in 41 (72%) of 57 adults with the retinal degeneration retinitis pigmentosa, while antibody responses to five other common neurotropic viruses were normal. However, these patients lacked clinical signs of active RV infection or known recent RV exposure, and 56 lacked IgM anti-RV antibody. Unusual relative percentages of IgG antibody to RV structural proteins compared with those of controls were found in patients' sera by radioimmunoprecipitation assay. For retinitis pigmentosa patients, percentage of RV envelope glycoprotein E1 antibody was similar to, of RV envelope glycoprotein E2 antibody was greater than, and of antibody to RV nucleocapsid C protein was lower than control percentages. Abnormal immunity to RV was also suggested by a lack of increased proliferation of lymphocytes to RV antigen despite elevated anti-RV antibody in patients with retinitis pigmentosa. Not associated with age or particular genetic pattern, these divergences from normal immunity suggest an unusual association between RV proteins and retinitis pigmentosa.

An expanded library of murine monoclonal antibodies (MAbs) was generated by infecting BALB/C mice with the Therien strain of rubella virus (RV) and selecting secreting hybrids by enzyme-linked immunosorbent assay (ELISA) using purified virion targets. A panel of plasmids containing specified RV cDNA fragments was also constructed by using a variety of strategies with pGE374- and pGE374-derived expression vectors. Hybrid RecA-RV-beta-galactosidase (LacZ)- or RecA-RV-truncated LacZ-containing proteins collectively representing the entire open reading frame of the structural proteins of RV were overexpressed in Escherichia coli. Bacterial lysates were then probed by ELISA with selected MAbs and by immunoblot following separation by electrophoresis under denaturing conditions. With this approach, MAbs that appeared to react with linear determinants defined epitopes localized within the following domains: MAbs C-1, C-2, and C-8 bind epitopes within the predicted amino-terminal 21 amino acids of the capsid region C9 to C29; MAb C-9 binds to a domain bounded by C64 and C97; MAbs E2-1 through E2-6 bind to the E2 glycoprotein backbone region from E2(1) to E2(115); MAbs E1-18 and E1-20 bind to the E1 glycoprotein region from E1(202) to E1(283). MAb E1-18 neutralizes RV infectivity; MAb E1-20 neutralizes infectivity and modestly inhibits hemagglutination. Analyses with selected synthetic peptides have confirmed several of the molecular domains deduced with the expressed proteins. These plasmid constructions and peptides have proven useful in beginning to unravel the molecular organization of several antigenic sites of this human pathogen.