Bacterial Cellulose Applied in Wound Dressing Materials: Production and Functional Modification - A Review.

In recent years, the development of new type wound dressings has gradually attracted more attention. Bacterial cellulose (BC) is a natural polymer material with various unique properties, such as ultrafine 3D nanonetwork structure, high water retention capacity, and biocompatibility. These properties allow BC to be used independently or in combination with different components (such as biopolymers and nanoparticles) to achieve diverse effects. This means that BC has great potential as a wound dressing. However, systematic summaries for the production and commercial application of BC-based wound dressings are still lacking. Therefore, this review provides a detailed introduction to the production fermentation process of BC, including various production strains and their biosynthetic mechanisms. Subsequently, with regard to the functional deficiencies of bacterial cellulose as a wound dressing, recent research progress in this area is enumerated. Finally, prospects are discussed for the low-cost production and high-value-added product development of BC-based wound dressings.

Activation of Stimulation of Interferon Genes (STING) Signal and Cancer Immunotherapy.

Stimulator of interferon gene (STING), an intracellular receptor in the endoplasmic reticulum, could induce the production of cytokines such as type I interferon (IFN) by activating the cGAS-STING signal pathway. In recent years, activation of STING has shown great potential to enhance anti-tumor immunity and reshape the tumor microenvironment, which is expected to be used in tumor immunotherapy. A number of STING agonists have demonstrated promising biological activity and showed excellent synergistic anti-tumor effects in combination with other cancer therapies in preclinical studies and some clinical trials. The combination of STING agonists and ICI also showed a potent effect in improving anti-tumor immunity. In this review, we introduce the cGAS-STING signaling pathway and its effect in tumor immunity and discuss the recent strategies of activation of the STING signaling pathway and its research progress in tumor immunotherapy.

Human IL-17 and TNF-α Additively or Synergistically Regulate the Expression of Proinflammatory Genes, Coagulation-Related Genes, and Tight Junction Genes in Porcine Aortic Endothelial Cells.

Immune rejection is the major limitation for porcine xenograft survival in primate recipients. Proinflammatory cytokines play important roles in immune rejection and have been found to mediate the pathological effects in various clinical and experimental transplantation trials. IL-17 and TNF-α play critical pathological roles in immune disorders, such as psoriasis and rheumatoid arthritis. However, the pathological roles of human IL-17 (hIL-17) and human TNF-α (hTNF-α) in xenotransplantation remain unclear. Here we found that hIL-17 and hTNF-α additively or synergistically regulate the expression of 697 genes in porcine aortic endothelial cells (PAECs). Overall, 415 genes were found to be synergistically regulated, while 282 genes were found to be additively regulated. Among these, 315 genes were upregulated and 382 genes were downregulated in PAECs. Furthermore, we found that hIL-17 and hTNF-α additively or synergistically induced the expression of various proinflammatory cytokines and chemokines (e.g., IL1α, IL6, and CXCL8) and decreased the expression of certain anti-inflammatory genes (e.g., IL10). Moreover, hIL-17 plus hTNF-α increased the expression of IL1R1 and IL6ST, receptors for IL1 and IL6, respectively, and decreased anti-inflammatory gene receptor expression (IL10R). hIL-17 and hTNF-α synergistically or additively induced CXCL8 and CCL2 expression and consequently promoted primary human neutrophil and human leukemia monocytic cell migration, respectively. In addition, hIL-17 and hTNF-α induced pro-coagulation gene (SERPINB2 and F3) expression and decreased anti-coagulation gene (TFPI, THBS1, and THBD) expression. Additionally, hIL-17 and hTNF-α synergistically decreased occludin expression and consequently promoted human antibody-mediated complement-dependent cytotoxicity. Interestingly, hTNF-α increased swine leukocyte antigen (SLA) class I expression; however, hIL-17 decreased TNF-α-mediated SLA-I upregulation. We concluded that hIL-17 and hTNF-α likely promote the inflammatory response, coagulation cascade, and xenoantibody-mediated cell injury. Thus, blockade of hIL-17 and hTNF-α together might be beneficial for xenograft survival in recipients.

Salivary microbiota analysis of patients with membranous nephropathy.

The oral microbiota are closely related to human health. Nonetheless, to the best of our knowledge, their relationship with membranous nephropathy (MN) remains unstudied. The saliva microbiota collected from 22 patients with MN and 15 healthy controls were analyzed by next­generation sequencing, and bioinformatics analysis of the 16S ribosomal RNA gene was subsequently carried out. The Chao1 and Shannon indices in patients with MN were higher than those in healthy controls. Analysis of similarities revealed that the oral microbiota in the patient group were significantly different from those in the healthy controls. At the genus level, the abundance of Alloprevotella, Granulicatella, Prevotella, Streptococcus and Prevotella_7 was markedly higher in patients with MN than in healthy controls. Six operational taxonomic units (OTUs; OTU5, OTU28, OTU9, OTU15, OTU33 and OTU38) were found to be markedly correlated with the clinical factors creatinine, proteinuria in 24 h, estimated glomerular filtration rate and systolic blood pressure. A total of 28 Kyoto Encyclopedia of Genes and Genomes pathways were obtained from the significant OTUs. The oral microbiota of patients with MN were investigated and it was found that OTU5, OTU28, OTU9, OTU15, OTU33 and OTU38 may be used as biomarkers. The present findings may assist in the diagnosis of patients with MN.

The Lineage Differentiation and Dynamic Heterogeneity of Thymic Epithelial Cells During Thymus Organogenesis.

Although much progress has been made recently in revealing the heterogeneity of the thymic stromal components, the molecular programs of cell lineage divergency and temporal dynamics of thymic epithelial cell (TEC) development are largely elusive. Here, we constructed a single-cell transcriptional landscape of non-hematopoietic cells from mouse thymus spanning embryonic to adult stages, producing transcriptomes of 30,959 TECs. We resolved the transcriptional heterogeneity of developing TECs and highlighted the molecular nature of early TEC lineage determination and cortico-medullary thymic epithelial cell lineage divergency. We further characterized the differentiation dynamics of TECs by clarification of molecularly distinct cell states in the thymus developing trajectory. We also identified a population of Bpifa1+ Plet1+ mTECs that was preserved during thymus organogenesis and highly expressed tissue-resident adult stem cell markers. Finally, we highlighted the expression of Aire-dependent tissue-restricted antigens mainly in Aire+ Csn2+ mTECs and Spink5+ Dmkn+ mTECs in postnatal thymus. Overall, our data provided a comprehensive characterization of cell lineage differentiation, maturation, and temporal dynamics of thymic epithelial cells during thymus organogenesis.

Salt-inducible kinases inhibitor HG-9-91-01 targets RIPK3 kinase activity to alleviate necroptosis-mediated inflammatory injury.

Receptor-interacting protein kinase 3 (RIPK3) functions as a central regulator of necroptosis, mediating signaling transduction to activate pseudokinase mixed lineage kinase domain-like protein (MLKL) phosphorylation. Increasing evidences show that RIPK3 contributes to the pathologies of inflammatory diseases including multiple sclerosis, infection and colitis. Here, we identified a novel small molecular compound Salt-inducible Kinases (SIKs) inhibitor HG-9-91-01 inhibiting necroptosis by targeting RIPK3 kinase activity. We found that SIKs inhibitor HG-9-91-01 could block TNF- or Toll-like receptors (TLRs)-mediated necroptosis independent of SIKs. We revealed that HG-9-91-01 dramatically decreased cellular activation of RIPK3 and MLKL. Meanwhile, HG-9-91-01 inhibited the association of RIPK3 with MLKL and oligomerization of downstream MLKL. Interestingly, we found that HG-9-91-01 also trigger RIPK3-RIPK1-caspase 1-caspase 8-dependent apoptosis, which activated cleavage of GSDME leading to its dependent pyroptosis. Mechanistic studies revealed that SIKs inhibitor HG-9-91-01 directly inhibited RIPK3 kinase activity to block necroptosis and interacted with RIPK3 and recruited RIPK1 to activate caspases leading to cleave GSDME. Importantly, mice pretreated with HG-9-91-01 showed resistance to TNF-induced systemic inflammatory response syndrome. Consistently, HG-9-91-01 treatment protected mice against Staphylococcus aureus-mediated lung damage through targeting RIPK3 kinase activity. Overall, our results revealed that SIKs inhibitor HG-9-91-01 is a novel inhibitor of RIPK3 kinase and a potential therapeutic target for the treatment of necroptosis-mediated inflammatory diseases.

Dysregulated Microbiota-Driven Gasdermin D Activation Promotes Colitis Development by Mediating IL-18 Release.

The balance between gut microbiota and host is critical for maintaining host health. Although dysregulation of the gut microbiota triggers the development of various inflammatory diseases, including colitis, the molecular mechanism of microbiota-driven colitis development is largely unknown. Here, we found that gasdermin D (GSDMD) was activated during acute colitis. In the dextran sulfate sodium (DSS)-induced colitis model, compared to wild-type mice, Gsdmd-deficient mice had less colitis severity. Mechanistically, GSDMD expression in intestinal epithelial cells (IECs), but not infiltrating immune cells, was critical for GSDMD-mediated colitis progression. Moreover, commensal Escherichia coli (E. coli) largely overgrew during colitis, and then the dysregulated commensal E. coli mediated GSDMD activation. Furthermore, the activated GSDMD promoted the release of interleukin-18 (IL-18), but not the transcript or maturation level of IL-18, which in turn mediated goblet cell loss to induce colitis development. Thus, GSDMD promotes colitis development by mediating IL-18 release, and the microbiota can mediate colitis pathogenesis through regulation of GSDMD activation. Our results provide a potential molecular mechanism by which the microbiota-driven GSDMD activation contributes to colitis pathogenesis.

Discovery of novel serum metabolic biomarkers in patients with polycystic ovarian syndrome and premature ovarian failure.

Several widely recognized metabolites play a role in regulating the pathophysiological processes of various disorders. Nonetheless, the lack of effective biomarkers for the early diagnosis of polycystic ovarian syndrome (PCOS) and premature ovarian failure (POF) has led to the discovery of serum-based metabolic biomarkers for these disorders. We aimed to identify various differentially expressed metabolites (DEMs) through serum-based metabolic profiling in patients with PCOS and POF and in healthy individuals by using liquid chromatography-mass spectrometry analysis. Furthermore, heatmap clustering, correlation, and Z-score analyses were performed to identify the top DEMs. Kyoto Encyclopedia of Genes and Genomes enriched pathways of DEMs were determined using metabolite-based databases. Moreover, the clinical significance of these DEMs was evaluated on the basis of area under the receiver operating characteristic curve. Significantly dysregulated expressions of several metabolites were observed in the intergroup comparisons of the PCOS, POF, and healthy control groups. Furthermore, 6 DEMs were most frequently observed among the three groups. The expressions of these DEMs were not only directly correlated but also exhibited potential significance in patients with PCOS and POF. Novel metabolites with up/downregulated expressions can be discovered in patients with PCOS and POF using serum-based metabolomics; these metabolites show good diagnostic performance and can act as effective biomarkers for the early detection of PCOS and POF. Furthermore, these metabolites might be involved in the pathophysiological mechanisms of PCOS and POF via interplay with corresponding genes.

Silencing of long non-coding RNA NEAT1 improves Treg/Th17 imbalance in preeclampsia via the miR-485-5p/AIM2 axis.

T-regulatory (Treg)/T-helper 17 (Th17) imbalance is associated with preeclampsia (PE). Herein, we aimed to explore the effect and mechanism of lncRNA NEAT1 on the Treg/Th17 balance. The levels of nuclear enriched abundant transcript 1 (NEAT1), miR-485-5p, and absent in melanoma 2 (AIM2) in CD4+ T cells were determined using real-time quantitative polymerase chain reaction (RT-qPCR). Treg and Th17 cells were examined using flow cytometry. The relationship between miR-485-5p and NEAT1 or AIM2 was assessed using a dual-luciferase reporter assay. Pearson's correlation coefficient was used to analyze the correlation. All the data indicated that NEAT1 was upregulated in PE. The number of Treg cells decreased and was negatively related to NEAT1, whereas the number of Th17 cells increased and was positively related to NEAT1 in PE. Knockdown of NEAT1 increased the Treg cells and Treg/Th17 but decreased Th17 cells. Furthermore, NEAT1 sponges miR-485-5p to suppress the target AIM2 levels. Inhibition of miR-485-5p or upregulation of AIM2 abrogated the effect on Treg/Th17 balance induced by knockdown of NEAT1. In conclusion, silencing of NEAT1 promoted Treg/Th17 balance via the miR-485-5p/AIM2 axis in PE, suggesting that NEAT1 is a potential target for the treatment of PE.

Celecoxib alleviates pathological cardiac hypertrophy and fibrosis via M1-like macrophage infiltration in neonatal mice.

Cardiac hypertrophy is an adaptive response to all forms of heart disease, including hypertension, myocardial infarction, and cardiomyopathy. Cyclooxygenase-2 (COX-2) overexpression results in inflammatory response, cardiac cell apoptosis, and hypertrophy in adult heart after injury. However, immune response-mediated cardiac hypertrophy and fibrosis have not been well documented in injured neonatal heart. This study showed that cardiac hypertrophy and fibrosis are significantly attenuated in celecoxib (a selective COX-2 inhibitor)-treated P8 ICR mice after cryoinjury. Molecular and cellular profiling of immune response shows that celecoxib inhibits the production of cytokines and the expression of adhesion molecular genes, increases the recruitment of M1-like macrophage at wound site, and alleviates cardiac hypertrophy and fibrosis. Furthermore, celecoxib administration improves cardiac function at 4 weeks after injury. These results demonstrate that COX-2 inhibition promotes the recruitment of M1-like macrophages during early wound healing, which may contribute to the suppression of cardiac hypertrophy and fibrosis after injury.

Tea polyphenol EGCG inhibited colorectal-cancer-cell proliferation and migration via downregulation of STAT3.

BACKGROUND: Green tea is a popular beverage worldwide and epigallocatechin-3-gallate (EGCG) is the most bioactive polyphenol in green tea. Our study aims to investigate the anti-proliferation and anti-migration effects of EGCG against colorectal-cancer SW480, SW620, and LS411N cells, and elucidate the underlying mechanism. METHODS: The in vitro anti-proliferation and anti-migration effects of EGCG against colon-cancer cells were evaluated using MTT, scratch-wound-healing, and transwell-migration assays. The effects of EGCG on apoptosis were assessed by Annexin V-FITC/PI double staining and JC-1 staining. Besides, Western blotting was employed to detect the protein-expression level and elucidate the underlying pathways. Real-time qPCR and dual-luciferase reporter assay were adopted to determine the mRNA level and promoter activity. RESULTS: Our results demonstrated that treatment with EGCG resulted in significant inhibition of cell proliferation by the induction of apoptosis. EGCG also inhibited SW480 cell migration in a dose-dependent manner as assessed by wound-healing and transwell-migration assays. Western blot confirmed that EGCG induced apoptosis by the activation of Caspase-3 and PARP. In addition, both STAT3 and phosphorylated STAT3 (p-STAT3) were downregulated significantly by EGCG in three selected colorectal-cancer cell lines. EGCG treatment also resulted in a significant decrease in Bcl-2, MCL-1, and Vimentin, and an increase in E-cadherin. When STAT3 was inhibited, EGCG showed no obvious effect on cell proliferation and migration. Further investigation by luciferase-reporter-activity assay showed that EGCG suppressed the promoter activity of STAT3 and downregulated the transcription of STAT3. CONCLUSION: Our study presents evidence on the anti-proliferation and anti-migration effects of EGCG against colorectal-cancer SW480, SW620, and LS411N cells by downregulating the expression of STAT3 and suggests that EGCG could be an effective and natural supplement for colon-cancer treatment.

A novel recessive mutation affecting DNAJB6a causes myofibrillar myopathy.

Mutations in the DNAJB6 gene have been identified as rare causes of myofibrillar myopathies. However, the underlying pathophysiologica mechanisms remain elusive. DNAJB6 has two known isoforms, including the nuclear isoform DNAJB6a and the cytoplasmic isoform DNAJB6b, which was thought to be the pathogenic isoform. Here, we report a novel recessive mutation c.695_699del (p. Val 232 Gly fs*7) in the DNAJB6 gene, associated with an apparently recessively inherited late onset distal myofibrillar myopathy in a Chinese family. Notably, the novel mutation localizes to exon 9 and uniquely encodes DNAJB6a. We further identified that this mutation decreases the mRNA and protein levels of DNAJB6a and results in an age-dependent recessive toxic effect on skeletal muscle in knock-in mice. Moreover, the mutant DNAJB6a showed a dose-dependent anti-aggregation effect on polyglutamine-containing proteins in vitro. Taking together, these findings reveal the pathogenic role of DNAJB6a insufficiency in myofibrillar myopathies and expand upon the molecular spectrum of DNAJB6 mutations.

Quantifying respiratory tract deposition of airborne graphene nanoplatelets: The impact of plate-like shape and folded structure.

The booming development of commercial products containing graphene nanoplatelets (GNPs) triggers growing concerns over their release into the air. Precise prediction of human respiratory system deposition of airborne GNPs, especially in alveolar region, is very important for inhalation exposure assessment. In this study, the pulmonary deposition of airborne GNPs was predicted by the multiple-path particle dosimetry (MPPD) model with consideration of GNPs plate-like shape and folded structure effect. Different equivalent diameters of GNPs were derived and utilized to describe different deposition mechanisms in the MPPD model. Both of small GNPs (geometric lateral size dg < 0.1 µm) and large GNPs (dg > 10 µm) had high deposition fractions in human respiratory system. The total deposition fractions for 0.1 and 30 µm GNPs were 41.6% and 75.6%, respectively. Most of the small GNPs deposited in the alveolar region, while the large GNPs deposited in the head airways. The aerodynamic diameter of GNPs was much smaller than the geometric lateral dimension due to the nanoscale thickness. For GNPs with geometric lateral size of 30 µm, the aerodynamic diameter was 2.98 µm. The small aerodynamic diameter of plate-like GNPs enabled deposition in the alveolar region, and folded GNPs had higher alveolar deposition than planar GNPs. Heavy breathing led to higher GNPs deposition fraction in head airways and lower deposition fractions in the alveolar region than resting breathing.

Immunosuppressive and metabolic agents that influence allo- and xenograft survival by in vivo expansion of T regulatory cells.

The transplanted organs or cells survive if the recipient receives adequate long-term immunosuppressive therapy. Immunosuppressive therapy combined with cell-based strategies (eg, regulatory T cell [Treg]-based therapy) promotes graft survival. A combination of Treg-based therapy and minimal or no immunosuppressive drug therapy would have the potential to minimize the risks of the complications and side effects of these drugs. Fortunately, some immunosuppressive and other agents not only impede the effector T cell response, but also help generate new CD4+ Tregs from conventional effector T cells. These agents include IL-2, TGF-ß, agents that block the CD40/CD40L costimulation pathway, mTOR inhibitors, and histone deacetylase inhibitors. Consequently, a state of relative unresponsiveness to the transplanted organ may be induced through the expansion of Tregs. We here review the effect of these various agents on expansion of CD4+ Tregs in allo- and xenotransplantation. The expansion of Tregs might allow a dose reduction of the standard immunosuppressive drugs.

Evaluation of Regeneration Processes for Filtering Facepiece Respirators in Terms of the Bacteria Inactivation Efficiency and Influences on Filtration Performance.

The regeneration of filtering facepiece respirators (FFRs) is of critical importance because of the severe shortage of FFRs during large-scale outbreaks of respiratory epidemics, such as COVID-19. Comprehensive experiments regarding FFR regeneration were performed in this study with model bacteria to illustrate the decontamination performance of the regeneration processes. The results showed that it is dangerous to use a contaminated FFR without any microbe inactivation treatment because the bacteria can live for more than 8 h. The filtration efficiency and surface electrostatic potential of 75% ethanol-treated FFRs were significantly reduced, and a most penetrating particle size of 200 nm was observed. Steam and microwave irradiation (MWI) showed promising decontamination performances, achieving 100% inactivation in 90 and 30 min, respectively. The filtration efficiencies of steam-treated FFRs for 50 and 100 nm particles decreased from 98.86% and 99.51% to 97.58% and 98.79%, respectively. Ultraviolet irradiation (UVI) effectively inactivated the surface bacteria with a short treatment of 5 min and did not affect the filtration performance. However, the UV dose reaching different layers of the FFP2 mask sample gradually decreased from the outermost layer to the innermost layer, while the model bacteria on the second and third layers could not be killed completely. UVI+MWI and steam were recommended to effectively decontaminate the used respirators and still maintain the respirators' filtration efficiency. The present work provides a comprehensive evaluation for FFR regeneration in terms of the filtration efficiencies for 50-500 nm particles, the electrostatic properties, mechanical properties, and decontamination effects.

EGCG Enhanced the Anti-tumor Effect of Doxorubicine in Bladder Cancer via NF-κB/MDM2/p53 Pathway.

Doxorubicin (DOX), the first-line chemotherapy for bladder cancer, usually induces side effects. We previously demonstrated that green tea polyphenol EGCG had potent anti-tumor effect in bladder cancer via down regulation of NF-κB. This study aimed to investigate the additive/synergistic effect EGCG and DOX against bladder cancer. Our results demonstrated that the combined use of DOX and EGCG inhibited T24 and SW780 cell proliferation. EGCG enhanced the apoptosis induction effect of DOX in both SW780 and T24 cells and resulted in significant differences. Besides, EGCG promoted the inhibitory effect of DOX against bladder cancer cell migration. In addition, the in vivo results demonstrated that DOX in combination with EGCG showed the most potent anti-tumor effects among DOX, EGCG and DOX+EGCG treatment groups. Further mechanistic studies determined that the combination of DOX and EGCG inhibited phosphorylated NF-κB and MDM2 expression, and up-regulated p53 expression in tumor, as assessed by western blot and immunohistochemistry. Western blot in SW780 cells also confirmed that the combined use of EGCG and DOX caused significant increase in p53, p21, and cleaved-PARP expression, and induced significant inhibition in phosphorylated NF-κB and MDM2. When NF-κB was inhibited, the expression of p53 and p-MDM2 were changed, and the combination of DOX and EGCG showed no obvious effect in transwell migration and cell viability. In conclusion, the novel application of chemotherapy DOX and EGCG demonstrated potent anti-tumor, anti-migration and anti-proliferation effects against bladder cancer. EGCG enhanced the anti-tumor effect of DOX in bladder cancer via NF-κB/MDM2/p53 pathway, suggesting the potential clinical application against bladder cancer patients.

Selective inhibition of cyclooxygenase-2 protects porcine aortic endothelial cells from human antibody-mediated complement-dependent cytotoxicity.

BACKGROUND: Cyclooxygenase-2 (COX-2) is an inducible enzyme with catalytic activity for biosynthesis of prostaglandins which are the key mediators of inflammation. COX-2 is also the therapeutic target for widely used non-steroidal anti-inflammatory drugs (NSAIDs). However, the involvement of COX-2 in xenotransplantation (eg, pig-to-non-human primate) remains poorly recognized. METHODS: We investigated the mechanisms that regulate COX-2 expression and the effects of COX-2 on porcine aortic endothelial cell (PAEC) viability using in vitro pig-to-primate xenotransplantation model and in vivo pig-to-mouse cellular transplant model. Regulation of COX-2 expression was assessed by real-time quantitative polymerase chain reaction (qPCR) and Western blotting. The effects of inhibition or downregulation of COX-2 on PAEC viability were assessed by propidium iodide (PI)-Annexin V staining and Cell Counting Kit-8 assay. RESULTS: Human serum triggered robust COX-2 expression in PAECs in a dose- and time-dependent manner. Induction of COX-2 expression by human serum was partially through activation of both canonical and non-canonical nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κb) signaling and increasing intracellular calcium. Cytokines like tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL-1ß), IL-17, were able to induce COX-2 expression. Selective inhibition of COX-2 by celecoxib dramatically decreased PAEC death in vitro and in vivo as defined by propidium iodide (PI)-Annexin V staining. Consistently, downregulation of COX-2 expression by NF-κb inhibitors or calcium chelator BAPTA decreased human serum-induced PAEC death as well. Silencing of COX-2 expression by small interfering RNA (siRNA) protected PAEC viability when transplanted under kidney capsule of C57BL/6 mice. CONCLUSIONS: Taken together, our data suggest that COX-2 is highly induced in PAECs by xenogenic serum and associated with human antibody-mediated complement-dependent cytotoxicity. COX-2 might be a potential therapeutic target to improve xenotransplantation.

Hypoxia-inducible factor-prolyl hydroxylase inhibitor ameliorates myopathy in a mouse model of chronic kidney disease.

Muscle wasting and diminished physical performance contribute to the morbidity and mortality of chronic kidney disease (CKD), for which no curative therapy exists. Accumulating evidence indicates that impaired angiogenesis occurs in the muscles of CKD models. Therefore, proangiogenesis therapy is considered a potentially effective strategy for limiting CKD-associated myopathy. Hypoxia-inducible factor (HIF)-prolyl hydroxylase inhibitor (HIF-PHI) stabilizes HIF and enhances muscle angiogenesis during acute ischemia; however, little evidence was available from CKD models. Here, we assessed whether pharmacological activation of HIF by MK-8617 (MK), a novel orally active HIF-PHI, improves CKD-associated myopathy. Mice were divided into sham or CKD groups, and CKD mice were subdivided into CKD + vehicle or MK treatment groups (1.5, 5, or 12.5 mg/kg for 12 wk). In CKD mice, skeletal muscle mass, mitochondrial amount, and exercise capacity decreased compared with sham mice. Compared with the CKD + vehicle group, low (1.5 mg/kg) and medium (5 mg/kg) doses of MK, but not the high dose (12.5 mg/kg), significantly restored these changes and was accompanied by incremental increases in HIF-1α. Furthermore, increased capillary density and area were observed in a MK dose-dependent manner, which is likely related to an improved VEGF response in the skeletal muscle of CKD mice. In addition, macrophage and proinflammatory cytokines, including monocyte chemoattractant protein 1, TNF-α, and IL-6, significantly increased in the high-dose MK group. These results indicate that HIF-PHI provides a potential therapeutic strategy to improve CKD-associated myopathy.

Isolation and Culture of Primary Aortic Endothelial Cells from Miniature Pigs.

Xenotransplantation is a promising way to resolve the shortage of human organs for patients with end-stage organ failure, and the pig is considered as a suitable organ source. Immune rejection and coagulation are two major hurdles for the success of xenotransplantation. Vascular endothelial cell (EC) injury and dysfunction are important for the development of the inflammation and coagulation responses in xenotransplantation. Thus, isolation of porcine aortic endothelial cells (pAECs) is necessary for investigating the immune rejection and coagulation responses. Here, we have developed a simple enzymatic approach for the isolation, characterization, and expansion of highly purified pAECs from miniature pigs. First, the miniature pig was anaesthetized with ketamine, and a length of aorta was excised. Second, the endothelial surface of aorta was exposed to 0.005% collagenase IV digestive solution for 15 min. Third, the endothelial surface of the aorta was scraped in only one direction with a cell scraper (<10 times), and was not compressed during the process of scraping. Finally, the isolated pAECs of Day 3, and after passage 1 and passage 2, were identified by flow cytometry with an anti-CD31 antibody. The percentages of CD31-positive cells were 97.4% ± 1.2%, 94.4% ± 1.1%, and 92.4% ± 1.7% (mean ± SD), respectively. The concentration of Collagenase IV, the digestive time, the direction, and frequency and intensity of scraping are critical for decreasing fibroblast contamination and obtaining high-purity and a large number of ECs. In conclusion, our enzymatic method is a highly-effctive method for isolating ECs from the miniature pig aorta, and the cells can be expanded in vitro to investigate the immune and coagulation responses in xenotransplantation.

Salivary microbial analysis of Chinese patients with immunoglobulin A nephropathy.

Microbiota plays an important role in immunoglobulin A (IgA) nephropathy (IgAN); however, the pathogenesis, early diagnosis, and treatment of IgAN remain unclear. The aim of the present study was to develop a preliminary model based on saliva­specific microbes and clinical indicators to facilitate the early diagnosis of IgAN and obtain insights into its treatment. The microbial profile of the saliva of 28 IgAN patients and 25 healthy control subjects was investigated using high­throughput sequencing and bioinformatics analyses of the V4 region in microbial 16S rRNA genes. IgAN patients and healthy subjects did not differ significantly in α­diversity indices (Chao1 and Shannon index) or phylum composition. At the genus level, however, Granulicatella was significantly less abundant in healthy individuals than in IgAN patients, while Prevotella and Veillonella were significantly more abundant in the healthy subjects than in IgAN patients (P<0.05 and P<0.01, respectively). Correlation analysis between biochemical indicators and operational taxonomic units (OTUs) revealed that the glomerular filtration rate was positively correlated with OTU86 and OTU287 at P<0.05, positively correlated with OTU165 at P<0.001, and negatively correlated with OTU455 at P<0.05. The serum creatinine index was negatively correlated with OTU287 at P<0.05 and negatively correlated with OTU165 at P<0.001. The pathological changes were positively correlated with OTU255 at P<0.05, OTU200 at P<0.01, and OTU455 and OTU75 at P<0.001, and negatively correlated with OTU86, OTU287, and OTU788 at P<0.05 and with OTU165 at P<0.01. The differences between Chinese IgAN patients and healthy subjects in terms of OTUs and biochemical indicators were analyzed and a mathematical model to facilitate the clinical diagnosis of IgAN was established.