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To explore the feasibility and safety of biomaterials for posterior scleral reinforcement (PSR) in rabbits. Decellularization and genipin crosslink were applied to the fresh bovine pericardium and porcine endocranium, and then mechanical properties, suture retention strength, and stability were tested. PSR operation was performed on 24 rabbit eyes using treated biological materials. Ophthalmic examination was performed regularly before and after PSR operation (1 week, 1 month, 3 months, 6 months). To evaluate the effectiveness, A ultrasound, diopter, and optical coherence tomography were conducted. General condition, fundus photograph, and pathological examination were recorded to evaluate the safety. Compared with genipin crosslinked bovine pericardium (Gen-BP) (21.29 ± 13.29 Mpa), genipin crosslinked porcine endocranium (Gen-PE) (34.85 ± 3.67 Mpa, P < 0.01) showed a closer elastic modulus to that of genipin crosslinked human sclera. There were no complications or toxic reactions directly related to the materials. Capillary hyperplasia, inflammatory cell infiltration, and collagen fiber deposition were observed, and the content of type I collagen fibers increased after PSR. Overall, the choroidal thickness of treated eyes was significantly thickened at different time points after PSR, which were 96.84 ± 21.08 μm, 96.72 ± 22.00 μm, 90.90 ± 16.57 μm, 97.28 ± 14.74 μm, respectively. The Gen-PE group showed changes that were almost consistent with the overall data. Gen-BP and Gen-PE are safe biological materials for PSR. The Gen-PE group demonstrated more significant advantages over the Gen-BP group in terms of material properties., (© 2024 IOP Publishing Ltd.)

Background: Streptococcus pneumoniae (Spn) has been a leading cause of bacterial meningitis in children. The most recent estimation of the global burden of Spn meningitis indicates a positive trajectory in eliminating Spn through the implementation of pneumococcal conjugate vaccines. However, continuous monitoring and assessment of the disease burden are necessary due to the evidence of serotype replacement, antibiotic resistance, and the impact of the recent COVID-19 pandemic., Objective: The aim of this systematic review is to provide an updated and focused assessment of the global and regional burden of Spn meningitis in children, which can guide policies and strategies to reduce the disease burden., Methods: Population-based studies published from January 1, 2000, to January 1, 2022, were preliminarily searched from the electronic databases PubMed, Embase, Global Health (CABI), and CINAHL Plus without any language restrictions. Studies were included if they reported the incidence, prevalence, mortality, or case-fatality ratio (CFR) for Spn meningitis in children aged 0-4 years; meningitis was confirmed by cerebrospinal fluid culture; the study period was a minimum of 1 year; the number of reported cases was at least 10; and the study had no methodological ambiguities. The article screening process follows the PRISMA (Preferred Reporting Items for Systematic reviews and Meta-Analyses) guidelines. Characteristics including study period, setting, World Health Organization region, income level, vaccination information, and participant data (age, number of cases, deaths, sequelae, and risk factors) will be extracted from the included studies. Search results will be updated and incorporated into our review prior to finalizing the extraction of data. Generalized linear mixed models meta-analysis will be performed to estimate the pooled incidence and CFR. We will further assess the risk of bias and heterogeneity, and will perform subgroup and sensitivity analyses to provide a meaningful interpretation of the current burden and literature for pneumococcal meningitis., Results: Our preliminary search in December 2021 yielded 9295 articles. Out of 275 studies that were assessed with our eligibility criteria, 117 articles were included. Data extraction and analysis are expected to be complete by January 2025. We plan to publish the results from the full study, including an updated search in 2024, by March 2025., Conclusions: Given that the major burden of Spn meningitis affects children under the age of 5 years, this systematic review will provide a thorough understanding of the global burden of Spn meningitis in this vulnerable population over a span of 2 decades. Insights into incidence trends, geospatial distribution, risk factors, and sequelae will be valuable for stakeholders, policy makers, and the academic community. This information will aid in the ongoing monitoring of the disease and in enhancing targeted vaccine programs to further mitigate the impact of the disease on children worldwide., Trial Registration: PROSPERO CRD42021293110; https://tinyurl.com/kc3j5k4m., International Registered Report Identifier (irrid): DERR1-10.2196/50678., (©Jay J Park, Jakov Tiefenbach, Mohammed Ma'arij Anwar, Sandhya Narayanan, Beatrice Ope, Selene Seo Bin Han, Boni Maxime Ale, Davies Adeloye, Igor Rudan. Originally published in JMIR Research Protocols (https://www.researchprotocols.org), 16.07.2024.)

Translation is a crucial process during plant growth and morphogenesis. In grapevine ( Vitis vinifera L.), many transcripts can be detected by RNA sequencing; however, their translational regulation is still largely unknown, and a great number of translation products have not yet been identified. Here, ribosome footprint sequencing was carried out to reveal the translational profile of RNAs in grapevine. A total of 8291 detected transcripts were divided into four parts, including the coding, untranslated regions (UTR), intron, and intergenic regions, and the 26 nt ribosome-protected fragments (RPFs) showed a 3 nt periodic distribution. Furthermore, the predicted proteins were identified and classified by GO analysis. More importantly, 7 heat shock-binding proteins were found to be involved in molecular chaperone DNA J families participating in abiotic stress responses. These 7 proteins have different expression patterns in grape tissues; one of them was significantly upregulated by heat stress according to bioinformatics research and was identified as DNA JA6. The subcellular localization results showed that VvDNA JA6 and VvHSP70 were both localized on the cell membrane. Therefore, we speculate that DNA JA6 may interact with HSP70. In addition, overexpression of VvDNA JA6 and VvHSP70 , reduced the malondialdehyde (MDA) content, improved the antioxidant enzyme activity of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), increased the content of proline, an osmolyte substance, and affected the expression of the high-temperature marker genes VvHsfB1, VvHsfB2A, VvHsfC and VvHSP100 . In summary, our study proved that VvDNA JA6 and the heat shock protein VvHSP70 play a positive role in the response to heat stress. This study lays a foundation for further exploring the balance between gene expression and protein translation in grapevine under heat stress., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhen, Dongying, Yue, Lipeng, Jingjing, Minying, Yuanyuan, Juan, Shiren, Yi, Bin and Chao.)

Background: Patients with severe aplastic anemia (SAA) frequently present with inflammatory episodes, and during flared inflammatory episodes, hematopoietic function is further exacerbated. The gastrointestinal tract is the most common site for infectious and inflammatory diseases, and its structural and functional features confer on it the most potent capacity to affect hematopoietic and immune functions. Computed tomography (CT) is a readily accessible approach to provide highly useful information in detecting morphological changes and guiding further work-ups., Aim: To explore CT imaging presentations of gut inflammatory damage in adult SAA patients during inflammatory episodes., Methods: We retrospectively evaluated the abdominal CT imaging presentations of 17 hospitalized adult patients with SAA in search of the inflammatory niche when they presented with systemic inflammatory stress and exacerbated hematopoietic function. In this descriptive manuscript, the characteristic images that suggested the presence of gastrointestinal inflammatory damage and related imaging presentations of individual patients were enumerated, analyzed and described., Results: All eligible patients with SAA had CT imaging abnormalities that suggested the presence of an impaired intestinal barrier and increased epithelial permeability. The inflammatory damages were concurrently present in the small intestine, the ileocecal region and the large intestines. Some readily identified imaging signs, such as bowel wall thickening with mural stratification ("water holo sign", "fat holo sign", intramural gas and subserosal pneumatosis) and mesenteric fat proliferation (fat stranding and "creeping fat sign"), fibrotic bowel wall thickening, "balloon sign", rugged colonic configuration, heterogeneity in the bowel wall texture, and adhered and clustered small bowel loop (including various patterns of "abdominal cocoon"), occurred at a high incidence, which suggested that the damaged gastrointestinal tract is a common inflammatory niche responsible for the systemic inflammatory stresses and the exacerbated hematopoietic failure in patients with SAA. Particularly, the "fat holo sign" was present in 7 patients, a rugged colonic configuration was present in 10 patients, the adhesive bowel loop was present in 15 patients, and extraintestinal manifestations suggestive of tuberculosis infections were present in 5 patients. According to the imaging features, a suggestive diagnosis of Crohn's disease was made in 5 patients, ulcerative colitis in 1 patient, chronic periappendiceal abscess in 1 patient, and tuberculosis infection in 5 patients. Other patients were diagnosed with chronic enteroclolitis with acutely aggravated inflammatory damage., Conclusion: Patients with SAA had CT imaging patterns that suggested the presence of active chronic inflammatory conditions and aggravated inflammatory damage during flared inflammatory episodes., Competing Interests: Conflict-of-interest statement: The authors have no conflicts of interest to declare that are relevant to the content of this article., (©The Author(s) 2023. Published by Baishideng Publishing Group Inc. All rights reserved.)

The process parameters and sludge properties of an in-situ sludge ozone-reduction system were investigated under different ozone dosages and sludge ages. Subsequently, 75 mg·g -1 (as O 3 /MLVSS) was selected as the appropriate ozone dosage to satisfy the wastewater treatment capacity and in-situ sludge ozone-reduction. The calibration coefficient results of the sludge yield formula indicated that Y h was reduced from 0.331 g·g -1 to 0.326 g·g -1 (to MLVSS/COD m) by ozone treatment and K d was increased from 0.046 d -1 to 0.050 d -1 at 75 mg·g -1 . The effluent quality of the SBR system was satisfactory when the sludge age was 10 d. The ozone dosage of 75 mg·g -1 and sludge age of 10 d were selected as the appropriate process conditions, at which the excess sludge was reduced by 12%. The high-throughput sequencing results concluded that the microorganisms in the excess sludge after ozonation were different in phylum and genus. After ozone treatment, the abundance of Bacteroidetes increased by 1.2 times and the relative abundance of Proteobacteria with nitrification and denitrification ability decreased from 24% to 18%. The reduction in the abundance of nitrobacteria affected the denitrification capacity of the sewage treatment system, but the total effluent nitrogen still met the I B discharge standard of pollutants for municipal wastewater treatment plants. The relative abundance of Lactococcus increased from 0.4% to 21.6%. Simultaneously, the concentration of macromolecular organic substances in the EPS of the excess sludge increased from 40.6 mg·g -1 to 54.6 mg·g -1 , while the CST increased from 15 s to 17 s after ozone treatment. The zeta potential decreased from -10.04 mV to -15.20 mV and the SVI of the excess sludge increased from 54 mL·g -1 to 62 mL·g -1 , thereby indicating that the sedimentation performance and dewaterability were affected to some extent. However, the SS of the effluent and the solids content of the sludge cake after extraction did not change significantly, the system could still operate stably, and the subsequent dewaterability of the excess sludge was not significantly affected.

Male fertility is closely related to the normal function of the hypothalamicpituitary- testicular axis. The testis is an important male reproductive organ that secretes androgen and produces sperm through spermatogenesis. Spermatogenesis refers to the process by which spermatogonial stem cells (SSCs) produce highly differentiated spermatozoa and is divided into three stages: mitosis, meiosis and spermiogenesis. Spermatogenesis requires SSCs to strike a proper balance between self-renewal and differentiation and the commitment of spermatocytes to meiosis, which involves many molecules and signalling pathways. Abnormal gene expression or signal transduction in the hypothalamus and pituitary, but particularly in the testis, may lead to spermatogenic disorders and male infertility. The phosphoinositol-3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signalling pathway is involved in many stages of male reproduction, including the regulation of the hypothalamus-pituitarygonad (HPG) axis during spermatogenesis, the proliferation and differentiation of spermatogonia and somatic cells, and the regulation of sperm autophagy and testicular endocrine function in the presence of environmental pollutants, particularly endocrinedisrupting chemicals (EDCs). In the PI3K/AKT/mTOR signalling pathway, mTOR is considered the central integrator of several signals, regulating metabolism, cell growth and proliferation. In particular, mTOR plays an important role in the maintenance and differentiation of SSCs, as well as in regulating the redox balance and metabolic activity of Sertoli cells, which play an important role in nutritional support during spermatogenesis., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background: Colorectal cancer (CRC) is a leading cause of cancer-related deaths worldwide. Growing evidence supports a role for noncoding RNAs (ncRNAs) in CRC. In particular, they form competitive endogenous RNA (ceRNA) networks involved in the regulation of mRNA expression. However, the role of these networks in the pathogenesis of CRC is not fully understood. The aim of this study was to elucidate the role of circRNA/lncRNA-miRNA-mRNA systems in CRC pathogenesis based on the construction of a ceRNA network., Methods: RNA expression profiles were obtained from public datasets in the Gene Expression Omnibus (GEO) database and used for further analysis by online databases and tools., Results: In total, 245 circRNAs, 1,666 lncRNAs, 5 miRNAs, and 934 mRNAs were differentially expressed in CRC samples. Functional enrichment analysis identified altered biological functions related to the mRNAs in the ceRNA network, and it was found that the oxytocin signaling pathway was significantly enriched (P<0.05) in genes with differential expression in CRC. Additionally, we established a protein-protein interaction (PPI) network and identified 10 hub genes for the construction of circRNA/lncRNA-miRNA-hub gene regulatory modules., Conclusions: We identified several ncRNAs with a possible pathogenetic role in CRC and built a CRC-specific ceRNA network. The results of our study provide novel insights into the molecular events implicated in CRC., Competing Interests: Conflicts of Interest: Both authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/tcr-19-2973). The authors have no conflicts of interest to declare., (2020 Translational Cancer Research. All rights reserved.)

Aiming at low precision of a micro-electromechanical systems (MEMS) gyroscope, the virtual gyroscope technology is used to fuse the data of several MEMS gyroscopes. In traditional virtual gyroscope technology, the angular acceleration of maneuver is modeled as white noise, which leads to low fusion accuracy in the dynamic state. So, the "current" probability density model is introduced to model the maneuvering angular rate accurately. The one-step predictive value of angular acceleration is approximated to the mean value of the current angular acceleration. And the variance of the angular acceleration is modeled by the one-step predictive value of angular acceleration. So, the "current" probability density model is used to realize the adaptive adjustment of the mean and variance of maneuvering angular acceleration, which improves the modeling accuracy of the maneuvering angular rate and the compensation accuracy of virtual gyroscope technology. The improved virtual gyroscope fusion method is verified in a low frequency, high frequency maneuver test. The experimental results show that the 1 σ standard deviation of the improved virtual gyroscope fusion error is 6.53 times lower than that of the single gyroscope static drift at low frequency and 2.04 times lower than that of the single gyroscope static drift at high frequency.

A series of novel Fe-Cd co-doped ZnO nanoparticle based photocatalysts are successfully synthesized by sol-gel route and characterized using scanning electron microscopy (SEM), energy dispersive X-ray emission (EDX), transmission electron microscopy (TEM), X-ray diffraction (XRD), UV-Vis spectroscopy, X-ray photoelectron spectroscopy (XPS), and Brunauer-Emmett-Teller (BET) techniques. The photocatalytic activity of ZnO nanoparticles doped with various atomic weight fraction of Fe and Cd has been investigated under visible light irradiation using the Methylene Blue and Rhodamine B dye in aqueous solution. The FeCd (2%):ZnO (ZFC-1) exhibit the highest photocatalytic activity in terms of rate constant as K MB  = 0.01153 min -1 and K RhB  = 0.00916 min -1 ). Further, the re-usability of the ZFC-1 photocatalyst is studied which confirms that it can be reused up to five times with nearly negligible loss of the photocatalytic efficiency. Moreover, the role of photoactive species investigated using a radical scavenger technique. The present investigations show that the doping concentration plays significant role in photocatalytic performance. The visible light absorption shown by Fe-Cd co-doped ZnO nanoparticles is much higher than that of undoped body probably due to co-doping, and the charge carrier recombination is decreased effectively which yields a higher photocatalytic performance. The mechanism for the enhancement of photocatalytic activity under visible light irradiation is also proposed.

We report an aerosol-based approach to study the thermal stability of metal-organic frameworks (MOFs) for gas-phase synthesis of MOF-based hybrid nanostructures used for highly active catalysis. Temperature-programmed electrospray-differential mobility analysis (TP-ES-DMA) provides the characterization of temperature-dependent morphological change directly in the gas phase, and the results are shown to be highly correlated with the structural thermal stability of MOFs determined by the traditional measurements of porosity and crystallinity. The results show that MOFs underwent thermal decomposition via simultaneous disassembly and deaggregation. Trimeric Cr-based MIL-88B-NH 2 exhibited a higher temperature of decomposition ( T d ), 350 °C, than trimeric Fe-based MIL-88B-NH 2 , 250 °C. For UiO-66, a significant decrease of T d by ≈100 °C was observed by using amine-functionalized ligands in the MOF structure. Copper oxide nanocrystals were successfully encapsulated in the UiO-66 crystal (Cu x O@UiO-66) by using a gas-phase evaporation-induced self-assembly approach followed by a suitable thermal treatment below T d (i.e., determined by TP-ES-DMA). Cu x O@UiO-66 demonstrated a very high catalytic activity and stability to CO oxidation, showing at least a 3-time increase in CO conversion compared to the bare CuO nanoparticle samples. The study demonstrates a prototype methodology (1) to determine structural thermal stability of MOFs using a gas-phase electrophoretic method (TP-ES-DMA) and (2) to gas-phase synthesize CuO nanocrystals encapsulated in MOFs.

Modifying the interfacial conditions of magnetic layers by capping with overlayers can efficiently enhance the magnetic functionality of a material. However, the mechanisms responsible for this are closely related to the crystalline structure, compositional combinations, and interfacial quality, and are generally complex. In this contribution, we explored the use of Ag ultrathin overlayers on annealed . A method for preparing magnetic layers with different levels of enhanced magnetic anisotropy energy was developed. The method essentially involves simply modifying the contact area of the metallic/magnetic interface. A rougher interface results in a larger contact area between the Ag and Ni layers, resulting in an increase in magnetic anisotropy energy. Moreover, post-annealing treatments led to the segregation of Ni atoms, thus making the enhancement in the coercive force even more efficient. A model permits an understanding of the contact area and a strategy for enhancing the magnetic anisotropy energy and the coercive force was developed. Our approaches and the developed model promise to be helpful in terms of developing potential applications of ultrathin magnetic layers in the area of spintronics.

Human cytomegalovirus (HCMV) has been recognized as a cause of severe, sometimes life-threatening disease in congenitally infected newborns as well as in immunocompromised individuals. However, the molecular mechanisms of the host-virus interaction remain poorly understood. Here, we profiled the expression of mRNAs and long noncoding RNAs (lncRNAs) in THP-1 cells using the emerging RNA-seq to investigate the transcriptional changes during HCMV latent infection. At 4 days post HCMV infection, a total of 169,008,624 sequence reads and 180,616 transcripts were obtained, respectively. Of these transcripts, 1,354 noncoding genes and 12,952 protein-coding genes were observed in Refseq database. Differential gene expression analysis identified 2,153 differentially expressed genes (DEGs) between HCMV-infected and mock-infected THP-1 cells, including 1,098 up-regulated genes and 1,055 down-regulated genes. These regulated genes were involved in pathways of apoptosis, inflammatory response and cell cycle progression, all of which may be implicated in viral pathogenesis. In addition, 646 lncRNAs (208 known lncRNAs and 438 novel lncRNAs) were upregulated and 424 (140 known and 284 novel) were downregulated in infected THP-1 cells. These findings have provided a dynamic scenario of DE candidate genes and lncRNAs at the virus-host interface and clearly warrant further experimental investigation associated with HCMV infection., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Objectives: TNF-α has been proved to be an effective target in rheumatoid arthritis treatment. So far, all the commercialised TNF-α antagonists function as passive immunotherapy. The aim of this study was to design a complex which can trigger active immunisation and overcome self-tolerance to elicit antibodies against murine TNF-α., Methods: The complex (KLH-TNF) was chemically synthesised by linking a selected peptide TNFα(4-23) from murine soluble TNF-α to a carrier protein, keyhole limpet haemocyanin (KLH). We evaluated its safety and antibody eliciting performance. We also evaluated its disease-regulating ability on collagen-induced arthritis models. Furthermore, the immune cells responses were analysed by T cell proliferation assay and B cell memory experiments., Results: The complex was safe without cytotoxity. The anti-mTNF-α antibody titers of the KLH-TNF group were 400 times greater than the control groups (p<0.0001). The elicited antibodies could combine with soluble TNF-α. The antibody response was independent of autologous TNF-α and could be reinforced by booster immunisation. Moreover, the complex did not trigger T cell activation and B cell memory response against native TNF-α. In animal experiments, KLH-TNF immunized mice showed a lower arthritis score (p<0.001) and better weight gain (p<0.01). Histological evaluations showed milder inflammation and cartilage depletion., Conclusions: Active immunotherapy against cytokine TNF-α is feasible by conjugating cytokine peptide with carrier protein. The elicited antibodies could combine with the native TNF-α and inhibit its activity. Importantly, the antibody response is reversible and independent of autologous TNF-α.

A scaleless wing mutant of silkworm, Bombyx mori, has much fewer scales than wild type (WT). The scaleless phenotype was associated with tracheal system developmental deficiency and excessive apoptosis of scale cells. In this study, the wing discs proteins of WT and scaleless during pupation were studied using 2-DE and mass spectrometry. Of the 99 identified protein spots, four critical differentially expressed proteins between WT and scaleless were further verified using Q-PCR. At the first day of pupation (P0) in WT, imaginal disk growth factor (IDGF) was upregulated, whereas actin-depolymerizing factor 1 (ADF1) and profilin (PFN), which associated with cellular motility and cytoplasmic extension, were downregulated. We speculated their coaction counteracts the correct organization of the tracheal system in wing disc. Thiol peroxiredoxin (TPx) was upregulated in scaleless at P0, but its mRNA higher expression occurred in the day before pupation (S4). TPx could inhibit the formation of hydrogen peroxide, preventing the release of cytochrome C and activation of the caspase family protease. Its higher expression in scaleless was responsible for the apoptosis of scale cells delayed. The results provide further evidence that the scaleless phenotype was related to the tracheal system developmental deficiency and excessive apoptosis of scale cells., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Purpose: Oxidative stress plays an important role in the pathogenesis of various ocular diseases such as retinopathy, glaucoma, and age-related macular degeneration. Activating transcription factor 4 (ATF4) is induced by various stressors, including endoplasmic reticulum (ER) and oxidative stress, and ATF4 expression is regulated translationally through the PERK pathway of eIF2α phosphorylation. Transcriptional regulation of the ATF4 gene under oxidative stress was investigated in human papillomavirus 16 (HPV-16)-transformed retinal pigment epithelial ARPE-19/HPV-16 cells., Methods: Retinal pigment epithelial cells, trabecular meshwork cells, and corneal endothelial cells were treated with anoxia and thapsigargin (TG). Gene expression of ATF4 and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and transcription factors was investigated by Western blot analysis, reporter assays, chromatin immunoprecipitation (ChIP) assays, and small interfering (si)RNA strategies. Cellular sensitivity to oxidative stress was determined., Results: The expression of two transcriptional factors, ATF4 and Nrf2, was significantly induced by anoxia and TG. The Nrf2 regulator Keap1 was downregulated by anoxia. Downregulation of Nrf2 abolished ATF4 expression. On the other hand, downregulation of Keap1 enhanced the expression of both Nrf2 and ATF4. The promoter activity of ATF4 was transactivated by the co-transfection of Nrf2 expression plasmids and reduced by the transfection of Nrf2-specific siRNA. The ChIP assays demonstrated that Nrf2 bound to the promoter of the ATF4 gene. Nrf2 downregulation nearly abolished the ATF4 induction by anoxia and TG. Consistent with these findings, the promoter activity of ATF4 was augmented by treatment with TG, HCA, H(2)O(2), and anoxia. However, stress induction of ATF4 promoter activity was observed, even when a mutation was introduced into the antioxidant-responsive elements site. Furthermore, stress induction of the ATF4 promoter was completely abolished when the 5' untranslated region of the ATF4 gene was deleted. Downregulation of ATF4 rendered ARPE-19/HPV-16 cells sensitive to oxidative stress., Conclusions: These results suggest that the stress induction of ATF4 is significantly regulated transcriptionally through a Nrf2-dependent mechanism and may be a double-edged sword in the pathogenesis of various retinopathies.

Several different in vivo and in vitro bioassays are used to evaluate melanosome transfer efficacy from melanocytes to keratinocytes. However, these methods are complicated and time consuming. Here, we report on a simple, rapid, direct, and reliable in vitro method for observing the process of melanosome transfer from melanocytes to keratinocytes. First, we selected and tested a melanoma cell line RPMI-7951 that can normally synthesize melanin and transfer from mature melanosomes to keratinocytes in vitro. We cocultured these cells with a human ovarian teratoma transformed epidermal carcinoma cell line, which is also capable of accepting melanosomes transferred from melanocytes, as in normal keratinocytes. The cells were cocultured for 24-72 h and double labeled with FITC-conjugated antibody against the melanosome-associated protein TRP-1, and with Cy5-conjugated antibody against the keratinocyte-specific marker keratin 14. The cells were examined by fluorescence microscope and flow cytometry. Melanosome transfer from melanocytes to keratinocytes increased in a time-dependent manner. To verify the accessibility of this method, the melanosome transfer inhibitor, a serine protease inhibitor, 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride, and a melanosome transfer stimulator, alpha-melanocyte-stimulating hormone, were added. The serine protease inhibitor decreased melanosome transfer, and alpha-melanocyte-stimulating hormone increased melanosome transfer, in a dose-dependent manner. In conclusion, this is a simple, rapid, and effective model system to quantify the melanosome transfer efficacy from melanocytes to keratinocytes in vitro.

Previous studies on azoospermia or oligozoospermia induced by heat stress or high doses of testosterone mainly focused on germ cell apoptosis; no data regarding their possible effect on spermatogonia mitosis are available. We have established unilateral cryptorchid and testosterone undecanoate (TU)-treated monkey models and examined expression of P16(INK4a) in the testis to look at its possible role in azoospermia or oligozoospermia induced by the heat stress or the TU treatment. The results showed that both heat stress and TU were capable of inducing expression of P16(INK4a) mainly in spermatogonia and other types of germ cells as well as Sertoli cells at the later stage of germ cell apoptosis, namely on Day 10 after operation or on Day 60 after TU injection. It is, therefore, suggested for the first time that P16(INK4a) protein may inhibit the spermatogonia mitosis in the testis at the later stage of the germ cell apoptosis, resulting in arrest of spermatogenesis.