Molecular Mechanism of Yangshen Maidong Decoction in the Treatment of Chronic Heart Failure based on Network Pharmacology, Molecular Docking, and Molecular Dynamics Simulations.

Chronic heart failure (CHF) is a complex multifactorial clinical syndrome leading to abnormal cardiac structure and function. The severe form of this ailment is characterized by high disability, high mortality, and morbidity. Worldwide, 2-17% of patients die at first admission, of which 17-45% die within 1 year of admission and >50% within 5 years. Yangshen Maidong Decoction (YSMDD) is frequently used to treat the deficiency and pain of the heart. The specific mechanism of action of YSMDD in treating CHF, however, remains unclear. Therefore, a network pharmacology-based strategy combined with molecular docking and molecular dynamics simulations was employed to investigate the potential molecular mechanism of YSMDD against CHF. The effective components and their targets of YSMDD and related targets of CHF were predicted and screened based on the public database. The network pharmacology was used to explore the potential targets and possible pathways that involved in YSMDD treated CHF. Molecular docking and molecular dynamics simulations were performed to elucidate the binding affinity between the YSMDD and CHF targets. Screen results, 10 main active ingredients, and 6 key targets were acquired through network pharmacology analysis. Pathway enrichment analysis showed that intersectional targets associated pathways were enriched in the Prostate cancer pathway, Hepatitis B pathway, and C-type lectin receptor signaling pathways. Molecular docking and molecular dynamics simulations analysis suggested 5 critical active ingredients have high binding affinity to the 5 key targets. This research shows the multiple active components and molecular mechanisms of YSMDD in the treatment of CHF and offers resources and suggestions for future studies.

[Establishment and Evaluation Strategy of an in vitro Cell Model of Bone Marrow Microenvironment Injury in Mouse Acute Graft-Versus-Host Disease].

OBJECTIVE: To establish a mesenchymal stem cell（MSC）-based in vitro cell model for the evaluation of mouse bone marrow acute graft-versus-host disease (aGVHD). METHODS: Female C57BL/6N mice aged 6-8 weeks were used as bone marrow and lymphocyte donors, and female BALB/c mice aged 6-8 weeks were used as aGVHD recipients. The recipient mouse received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) in 6-8 hours post irradiation to establish a bone marrow transplantation (BMT) mouse model (n=20). In addition, the recipient mice received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) and spleen lymphocytes (2×106/mouse) in 6-8 hours post irradiation to establish a mouse aGVHD model (n=20). On the day 7 after modeling, the recipient mice were anesthetized and the blood was harvested post eyeball enucleation. The serum was collected by centrifugation. Mouse MSCs were isolated and cultured with the addition of 2%, 5%, and 10% recipient serum from BMT group or aGVHD group respectively. The colony-forming unit-fibroblast（CFU-F) experiment was performed to evaluate the potential effects of serums on the self-renewal ability of MSC. The expression of CD29 and CD105 of MSC was evaluated by immunofluorescence staining. In addition, the expression of self-renewal-related genes including Oct-4, Sox-2, and Nanog in MSC was detected by real-time fluorescence quantitative PCR（RT-qPCR). RESULTS: We successfully established an in vitro cell model that could mimic the bone marrow microenvironment damage of the mouse with aGVHD. CFU-F assay showed that, on day 7 after the culture, compared with the BMT group, MSC colony formation ability of aGVHD serum concentrations groups of 2% and 5% was significantly reduced （P < 0．05）; after the culture, at day 14, compared with the BMT group, MSC colony formation ability in different aGVHD serum concentration was significantly reduced （P < 0．05）. The immunofluorescence staining showed that, compared with the BMT group, the proportion of MSC surface molecules CD29+ and CD105+ cells was significantly dereased in the aGVHD serum concentration group (P < 0.05), the most significant difference was at a serum concentration of 10% （P < 0．001, P < 0.01）. The results of RT-qPCR detection showed that the expression of the MSC self-renewal-related genes Oct-4, Sox-2, and Nanog was decreased, the most significant difference was observed at an aGVHD serum concentration of 10% （P < 0.01,P < 0．001,P < 0．001）. CONCLUSION: By co-culturing different concentrations of mouse aGVHD serum and mouse MSC, we found that the addition of mouse aGVHD serum at different concentrations impaired the MSC self-renewal ability, which providing a new tool for the field of aGVHD bone marrow microenvironment damage.

Albiflorin ameliorates thioacetamide-induced hepatic fibrosis: The involvement of NURR1-mediated inflammatory signaling cascades in hepatic stellate cells activation.

Thioacetamide (TAA) within the liver generates hepatotoxic metabolites that can be induce hepatic fibrosis, similar to the clinical pathological features of chronic human liver disease. The potential protective effect of Albiflorin (ALB), a monoterpenoid glycoside found in Paeonia lactiflora Pall, against hepatic fibrosis was investigated. The mouse hepatic fibrosis model was induced with an intraperitoneal injection of TAA. Hepatic stellate cells (HSCs) were subjected to treatment with transforming growth factor-beta (TGF-ß), while lipopolysaccharide/adenosine triphosphate (LPS/ATP) was added to stimulate mouse peritoneal macrophages (MPMs), leading to the acquisition of conditioned medium. For TAA-treated mice, ALB reduced ALT, AST, HYP levels in serum or liver. The administration of ALB reduced histopathological abnormalities, and significantly regulated the expressions of nuclear receptor-related 1 protein (NURR1) and the P2X purinoceptor 7 receptor (P2×7r) in liver. ALB could suppress HSCs epithelial-mesenchymal transition (EMT), extracellular matrix (ECM) deposition, and pro-inflammatory factor level. ALB also remarkably up-regulated NURR1, inhibited P2×7r signaling pathway, and worked as working as C-DIM12, a NURR1 agonist. Moreover, deficiency of NURR1 in activated HSCs and Kupffer cells weakened the regulatory effect of ALB on P2×7r inhibition. NURR1-mediated inhibition of inflammatory contributed to the regulation of ALB ameliorates TAA-induced hepatic fibrosis, especially based on involving in the crosstalk of HSCs-macrophage. Therefore, ALB plays a significant part in the mitigation of TAA-induced hepatotoxicity this highlights the potential of ALB as a protective intervention for hepatic fibrosis.

Regulation of the Nur77-P2X7r Signaling Pathway by Nodakenin: A Potential Protective Function against Alcoholic Liver Disease.

Alcoholic liver disease (ALD) is the main factor that induces liver-related death worldwide and represents a common chronic hepatopathy resulting from binge or chronic alcohol consumption. This work focused on revealing the role and molecular mechanism of nodakenin (NK) in ALD associated with hepatic inflammation and lipid metabolism through the regulation of Nur77-P2X7r signaling. In this study, an ALD model was constructed through chronic feeding of Lieber-DeCarli control solution with or without NK treatment. Ethanol (EtOH) or NK was administered to AML-12 cells, after which Nur77 was silenced. HepG2 cells were exposed to ethanol (EtOH) and subsequently treated with recombinant Nur77 (rNur77). Mouse peritoneal macrophages (MPMs) were treated with lipopolysaccharide/adenosine triphosphate (LPS/ATP) and NK, resulting in the generation of conditioned media. In vivo, histopathological alterations were markedly alleviated by NK, accompanied by reductions in serum triglyceride (TG), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels and the modulation of Lipin-1, SREBP1, and Nur77 levels in comparison to the EtOH-exposed group (p < 0.001). Additionally, NK reduced the production of P2X7r and NLRP3. NK markedly upregulated Nur77, inhibited P2X7r and Lipin-1, and promoted the function of Cytosporone B, a Nur77 agonist (p < 0.001). Moreover, Nur77 deficiency weakened the regulatory effect of NK on P2X7r and Lipin-1 inhibition (p < 0.001). In NK-exposed MPMs, cleaved caspase-1 and mature IL-1ß expression decreased following LPS/ATP treatment (p < 0.001). NK also decreased inflammatory-factor production in primary hepatocytes stimulated with MPM supernatant. NK ameliorated ETOH-induced ALD through a reduction in inflammation and lipogenesis factors, which was likely related to Nur77 activation. Hence, NK is a potential therapeutic approach to ALD.

Microgel-based carriers enhance skeletal stem cell reprogramming towards immunomodulatory phenotype in osteoarthritic therapy.

Skeletal stem cells (SSC) have gained attentions as candidates for the treatment of osteoarthritis due to their osteochondrogenic capacity. However, the immunomodulatory properties of SSC, especially under delivery operations, have been largely ignored. In the study, we found that Pdpn+ and Grem1+ SSC subpopulations owned immunoregulatory potential, and the single-cell RNA sequencing (scRNA-seq) data suggested that the mechanical activation of microgel carriers on SSC induced the generation of Pdpn+Grem1+Ptgs2+ SSC subpopulation, which was potent at suppressing macrophage inflammation. The microgel carriers promoted the YAP nuclear translocation, and the activated YAP protein was necessary for the increased expression of Ptgs2 and PGE2 in microgels-delivered SSC, which further suppressed the expression of TNF-ɑ, IL-1ß and promoted the expression of IL-10 in macrophages. SSC delivered with microgels yielded better preventive effects on articular lesions and macrophage activation in osteoarthritic rats than SSC without microgels. Chemically blocking the YAP and Ptgs2 in microgels-delivered SSC partially abolished the enhanced protection on articular tissues and suppression on osteoarthritic macrophages. Moreover, microgel carriers significantly prolonged SSC retention time in vivo without increasing SSC implanting into osteoarthritic joints. Together, our study demonstrated that microgel carriers enhanced SSC reprogramming towards immunomodulatory phenotype to regulate macrophage phenotype transformation for effectively osteoarthritic therapy by promoting YAP protein translocation into nucleus. The study not only complement and perfect the immunological mechanisms of SSC-based therapy at the single-cell level, but also provide new insight for microgel carriers in stem cell-based therapy.

TNFAIP3 Derived from Skeletal Stem Cells Alleviated Rat Osteoarthritis by Inhibiting the Necroptosis of Subchondral Osteoblasts.

Recent investigations have shown that the necroptosis of tissue cells in joints is important in the development of osteoarthritis (OA). This study aimed to investigate the potential effects of exogenous skeletal stem cells (SSCs) on the necroptosis of subchondral osteoblasts in OA. Human SSCs and subchondral osteoblasts isolated from human tibia plateaus were used for Western blotting, real-time PCR, RNA sequencing, gene editing, and necroptosis detection assays. In addition, the rat anterior cruciate ligament transection OA model was used to evaluate the effects of SSCs on osteoblast necroptosis in vivo. The micro-CT and pathological data showed that intra-articular injections of SSCs significantly improved the microarchitecture of subchondral trabecular bones in OA rats. Additionally, SSCs inhibited the necroptosis of subchondral osteoblasts in OA rats and necroptotic cell models. The results of bulk RNA sequencing of SSCs stimulated or not by tumor necrosis factor α suggested a correlation of SSCs-derived tumor necrosis factor α-induced protein 3 (TNFAIP3) and cell necroptosis. Furthermore, TNFAIP3-derived from SSCs contributed to the inhibition of the subchondral osteoblast necroptosis in vivo and in vitro. Moreover, the intra-articular injections of TNFAIP3-overexpressing SSCs further improved the subchondral trabecular bone remodeling of OA rats. Thus, we report that TNFAIP3 from SSCs contributed to the suppression of the subchondral osteoblast necroptosis, which suggests that necroptotic subchondral osteoblasts in joints may be possible targets to treat OA by stem cell therapy.

Human osteoarthritic articular cartilage stem cells suppress osteoclasts and improve subchondral bone remodeling in experimental knee osteoarthritis partially by releasing TNFAIP3.

BACKGROUND: Though articular cartilage stem cell (ACSC)-based therapies have been demonstrated to be a promising option in the treatment of diseased joints, the wide variety of cell isolation, the unknown therapeutic targets, and the incomplete understanding of the interactions of ACSCs with diseased microenvironments have limited the applications of ACSCs. METHODS: In this study, the human ACSCs have been isolated from osteoarthritic articular cartilage by advantage of selection of anatomical location, the migratory property of the cells, and the combination of traumatic injury, mechanical stimuli and enzymatic digestion. The protective effects of ACSC infusion into osteoarthritis (OA) rat knees on osteochondral tissues were evaluated using micro-CT and pathological analyses. Moreover, the regulation of ACSCs on osteoarthritic osteoclasts and the underlying mechanisms in vivo and in vitro were explored by RNA-sequencing, pathological analyses and functional gain and loss experiments. The one-way ANOVA was used in multiple group data analysis. RESULTS: The ACSCs showed typical stem cell-like characteristics including colony formation and committed osteo-chondrogenic capacity. In addition, intra-articular injection into knee joints yielded significant improvement on the abnormal subchondral bone remodeling of osteoarthritic rats. Bioinformatic and functional analysis showed that ACSCs suppressed osteoarthritic osteoclasts formation, and inflammatory joint microenvironment augmented the inhibitory effects. Further explorations demonstrated that ACSC-derived tumor necrosis factor alpha-induced protein 3 (TNFAIP3) remarkably contributed to the inhibition on osteoarhtritic osteoclasts and the improvement of abnormal subchondral bone remodeling. CONCLUSION: In summary, we have reported an easy and reproducible human ACSC isolation strategy and revealed their effects on subchondral bone remodeling in OA rats by releasing TNFAIP3 and suppressing osteoclasts in a diseased microenvironment responsive manner.

Regulation of Nur77-TLR4/MyD88 signaling pathway is required for Ginsenoside Rc ameliorates hepatic fibrosis regression by deactivating hepatic stellate cells.

HSCs (hepatic stellate cells) contribute to the excessive extracellular matrix (ECM) deposition plays a key role in the progression of hepatic fibrosis. The present study focused on the hepatoprotective effect of Ginsenoside Rc (Rc), one of the protopanaxadiol type ginsenoside, which has contributed to reverse activated HSCs to improve hepatic fibrosis via regulating Nur77-TLR4/MyD88 signaling pathway. We established the hepatic fibrosis model by intraperitoneal injection of carbon tetrachloride (CCl4). And HSCs were stimulated with TGF-ß, followed by silencing of Nur77, and then incubated in Rc. Rc significantly alleviated histopathological changes, reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. Rc could upregulate the Nur77 and downregulate fibrosis markers in the liver of mice, including decreasing the expressions of α-SMA, Collagen-I, the ratio of TIMP-1/MMP-13. Rc significantly increased the expression of Nur77 and suppressed the production of ECM in HSCs. Rc inhibited TLR4 signaling pathway, consequently reversing the inflammatory response, including the production of MyD88, IRAK1, IRAK4 and IL-23. When Nur77 was knocked in TGF-ß-stimulated HSCs, TLR4 and α-SMA production were increased. Rc suppressed these activatory effects in Nur77 knockdown HSCs. Rc reduced inflammatory reaction by regulating the Nur77-TLR4 signaling pathway while suppressing the fibrogenesis suggesting, underscoring a promising approach of Rc for the treatment in hepatic fibrosis. Targeting Nur77-TLR4 signaling in HSCs would be the potential strategy for Rc against hepatic fibrosis.

Ginsenoside Rd, a natural production for attenuating fibrogenesis and inflammation in hepatic fibrosis by regulating the ERRα-mediated P2X7r pathway.

Ginseng, when used as a food and nutritional supplement, has the ability to regulate human immunity. Here, the potential anti-hepatic fibrosis effect of ginsenoside Rd (Rd), one of the protopanaxadiol types of ginsenoside, was investigated. We established a hepatic fibrosis model using intraperitoneal injection of thioacetamide (TAA) for five weeks in mice. In addition, an in vitro model was established by using TGF-ß to activate hepatic stellate cells (HSCs), treated with Rd and an estrogen-related receptor α (ERRα) inhibitor (XCT-790). The ERRα knockdown (shRNA-ERRα) of the primary mouse hepatocytes was used to establish hepatocyte injury by TGF-ß, and they were then incubated in Rd. The Rd significantly alleviated the histopathological changes, and reduced the serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. The Rd could upregulate the ERRα and downregulate the fibrosis markers in the livers of mice. In TAA-induced mice, the Rd inhibited the purinergic ligand-gated ion channel 7 receptor (P2X7r)-mediated NLRP3 inflammasome activation, consequently reversing the liver inflammatory response. The Rd significantly increased the expression of ERRα and suppressed the extracellular matrix (ECM) in the HSCs or primary hepatocytes. The Rd significantly decreased the P2X7r-mediated NLRP3 inflammasome activation, consequently reversing the inflammatory response, including the production of IL-1ß, IL-23 in the activated HSCs and primary hepatocytes. The Rd could ameliorate the damage of the hepatocytes and further inhibit the entry of IL-1ß and IL-18 into the extracellular matrix. The Rd reduced the inflammatory reaction by regulating the ERRα-P2X7r signaling pathway while suppressing the fibrogenesis, which suggests that the Rd can serve as a novel dietary supplement approach to combat hepatic fibrosis.

Single-cell transcriptomic analysis identifies a highly replicating Cd168+ skeletal stem/progenitor cell population in mouse long bones.

Skeletal stem/progenitor cells (SSPCs) are tissue-specific stem/progenitor cells localized within skeletons and contribute to bone development, homeostasis, and regeneration. However, the heterogeneity of SSPC populations in mouse long bones and their respective regenerative capacity remain to be further clarified. In this study, we perform integrated analysis using single-cell RNA sequencing (scRNA-seq) datasets of mouse hindlimb buds, postnatal long bones, and fractured long bones. Our analyses reveal the heterogeneity of osteochondrogenic lineage cells and recapitulate the developmental trajectories during mouse long bone growth. In addition, we identify a novel Cd168+ SSPC population with highly replicating capacity and osteochondrogenic potential in embryonic and postnatal long bones. Moreover, the Cd168+ SSPCs can contribute to newly formed skeletal tissues during fracture healing. Furthermore, the results of multicolor immunofluorescence show that Cd168+ SSPCs reside in the superficial zone of articular cartilage as well as in growth plates of postnatal mouse long bones. In summary, we identify a novel Cd168+ SSPC population with regenerative potential in mouse long bones, which adds to the knowledge of the tissue-specific stem cells in skeletons.

Characterization of Phenylalanine Ammonia Lyases from Lettuce (Lactuca sativa L.) as Robust Biocatalysts for the Production of d- and l-Amino Acids.

Phenylalanine ammonia lyase (PAL) catalyzes the reversible conversion of l-phenylalanine into the corresponding trans-cinnamic acid, providing a route to optically pure α-amino acids. We explored the catalytic function of all five PALs encoded in the genome of lettuce (Lactuca sativa L.) that are previously known to be involved in wound browning. All LsPALs were active toward l-phenylalanine in the ammonia elimination reaction and displayed maximum activity at 55-60 °C and pH 9.0-9.5. However, four of them, LsPAL1-LsPAL4, showed significantly higher activity and thermal stability than LsPAL5, as well as a broader substrate spectrum including some challenging substrates with steric demanding or electron-donating substituents. The best one LsPAL3 was subjected to the kinetic resolution of a panel of 21 rac-phenylalanine derivatives, as well as the ammonia addition of 21 cinnamic acid derivatives. It showed excellent enantioselectivity in most cases and significantly better activity than previously described PALs for a number of challenging non-natural substrates, demonstrating its great potential in biocatalysis.

[Establishment and Evaluation of Intestinal Injury Model of Mouse Acute Graft Versus Host Disease Based on An Organoid Technology].

OBJECTIVE: To establish an intestinal organoid model that mimic acute graft versus host disease (aGVHD) caused intestinal injuries by using aGVHD murine model serum and organoid culture system, and explore the changes of aGVHD intestine in vitro by advantage of organoid technology. METHODS: 20-22 g female C57BL/6 mice and 20-22 g female BALB/c mice were used as donors and recipients for bone marrow transplantation, respectively. Within 4-6 h after receiving a lethal dose (8.0 Gy) of γ ray total body irradiation, a total of 0.25 ml of murine derived bone marrow cells (1×107/mice, n=20) and spleen nucleated cells (5×106/mice, n=20) was infused to establish a mouse model of aGVHD (n=20). The aGVHD mice were anesthetized at the 7th day after transplantation, and the veinal blood was harvested by removing the eyeballs, and the serum was collected by centrifugation. The small intestinal crypts of healthy C57BL/6 mice were harvested and cultivated in 3D culture system that maintaining the growth and proliferation of intestinal stem cells in vitro. In our experiment, 5%, 10%, 20% proportions of aGVHD serum were respectively added into the organoid culture system for 3 days. The formation of small intestinal organoids were observed under an inverted microscope and the morphological characteristics of intestinal organoids in each groups were analyzed. For further evaluation, the aGVHD intestinal organoids were harvested and their pathological changes were observed. Combined with HE staining, intestinal organ morphology evaluation was performed. Combined with Alcian Blue staining, the secretion function of aGVHD intestinal organoids was observed. The distribution and changes of Lgr5+ and Clu+ intestinal stem cells in intestinal organoids were analyzed under the conditions of 5%, 10% and 20% serum concentrations by immunohistochemical stainings. RESULTS: The results of HE staining showed that the integrity of intestinal organoids in the 5% concentration serum group was better than that in the 10% and 20% groups. The 5% concentration serum group showed the highest number of organoids, the highest germination rate and the lowest pathological score among experimental groups, while the 20% group exhibited severe morphological destruction and almost no germination was observed, and the pathological score was the highest among all groups(t=3.668, 4.334,5.309,P<0.05). The results of Alican blue staining showed that the secretion function of intestinal organoids in serum culture of aGVHD in the 20% group was weaker than that of the 5% group and 10% of the organoids, and there was almost no goblet cells, and mucus was stainned in the 20% aGVHD serum group. The immunohistochemical results showed that the number of Lgr5+ cells of intestinal organoids in the 5% group was more than that of the intestinal organoids in the 10% aGVHD serum group and 20% aGVHD serum group. Almost no Clu+ cells were observed in the 5% group. The Lgr5+ cells in the 20% group were seriously injuried and can not be observed. The proportion of Clu+ cells in the 20% group significantly increased. CONCLUSION: The concentration of aGVHD serum in the culture system can affect the number and secretion function of intestinal organoids as well as the number of intestinal stem cells in organoids. The higher the serum concentration, the greater the risk of organoid injury, which reveal the characteristics of the formation and functional change of aGVHD intestinal organoids, and provide a novel tool for the study of intestinal injury in aGVHD.

Interpretation of guidelines for the prevention and treatment of chronic hepatitis B (2022 edition) / 临床肝胆病杂志

Guidelines for the prevention and treatment of chronic hepatitis B (2022 edition) are updated and revised based on the research advances in chronic hepatitis B virus infection in China and globally and the previous editions of the guidelines. This article introduces the updates in natural history and the noninvasive diagnosis and treatment of fibrosis. In particular, the guidelines further expand the indications for patients with chronic hepatitis B virus infection, clearly defines the selection of the population benefiting from interferon therapy, and strictly limits the standard of oral nucleos(t)ide analogues. Meanwhile, the guidelines also recommend more active treatment of patients with low-level viremia and children in the immune-tolerant phase. The new edition of the guidelines will provide an important basis for expanding the screening for hepatitis B virus infection, improving diagnostic rate, optimizing treatment regimens, and standardizing clinical management in China.

Clinical value of highly sensitive hepatitis B virus surface antigen and DNA assays / 中华检验医学杂志

For patients with chronic hepatitis B, the indications of antiviral treatment are gradually expanding, and the pursuit of various degrees of cure (partial cure, functional cure and complete cure) is consistently improving, which enhances the urgent clinical need for improving the detection sensitivity of hepatitis B virus (HBV) surface antigen and DNA. Based on the availability of commercial highly sensitive hepatitis B surface antigen and HBV DNA detection reagents, we summarized their applications in the diagnosis of HBV infection, their role on guiding selection of antiviral treatment agents and treatment plans, their prediction efficacy and indication for drug withdrawal, their role on monitoring therapy efficacy and the prediction of disease outcomes. Taken together, highly sensitive hepatitis B virus surface antigen and DNA assays and related detection technology play an important role in the whole management process of chronic HBV infection.

Clinical value of a novel biological mesh in laparoscopic inguinal hernia repair: a multicenter prospective randomized controlled study / 中华消化外科杂志

Objectives:To investigate the clinical value of a novel non-crosslinked biological mesh in laparoscopic inguinal hernia repair.Methods:The prospective randomized controlled study was conducted. The clinical data of 50 adult patients with unilateral inguinal hernia who were admitted to 3 medical centers, including Ruijin Hospital of Shanghai Jiaotong University School of Medicine et al, from September 2019 to March 2020 were selected. Based on random number table, patients were divided into two groups. Patients using the novel non-crosslinked biological mesh in repair surgery were divided into the experiment group and patients using the lightweight, micro-porous, partially absorbable synthetic mesh in repair surgery were divided into the control group. Observation indicators: (1) grouping situations of the enrolled patients; (2) endpoint of the study. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the t test. Measurement data with skewed distribution were represented as M(range), and comparison between groups was conducted using the non-parameter rank sum test. Count data were described as absolute numbers and (or) persentages, and comparison between groups was conducted using the chi-square test or Fisher exact probability. Comparison of ordinal data was conducted using the non-parameter rank sum test. Repeated measurement data were analyzed using the repeated ANOVA. Taking the recurrence rate of hernia as the basis of efficacy evaluation, according to the intention-to-treat analysis, the confidence interval method (Newcombe Wilson method) was used to conduct non-inferiority statistical analysis of the recurrence rate of hernia between the experiment group and the control group. If the upper limit of 95% confidence interval of the difference of recurrence rate of hernia between the experiment group and the control group is less than 10%, the experiment group is considered to be non-inferior to the control group. Results:(1) Grouping situations of the enrolled patients. A total of 50 adult patients with inguinal hernia were selected for eligibility. There were 44 males and 6 females, aged (60±15)years. All 50 patients were randomly divided into to the experiment group and the control group with 25 cases each. One patient in the control group was not followed up at postoperative month 2, and the rest of 49 patients completed all expected follow-up. No patient in the two groups fell off or were removed. (2) Endpoint of the study. ① The primary endpoint of study. The recurrence rate of hernia was 0 in the experiment group, versus 4%(1/25) in the control group, respectively, showing no significant difference between the two groups ( P>0.05). Results of non-inferiority statistical analysis showed that the 95% confidence interval of the difference of recurrence rate of hernia between the two groups was -19.54% to 9.72%, with the upper limit as 9.72%, which was less than 10%. ② The secondary endpoint of study. There were 2 patients in the control group occurred seroma at postoperative day 14, and none of the rest of patient in the two groups occurred seroma during the follow-up, showing no significant difference in the occurrence of seroma between the two groups ( P>0.05). There was 1 patient in the control group feeling discomfort or foreign body sensation in groin area at postoperative month 2, and none of the rest of patient in the two groups feeling discomfort or foreign body sensation in groin area during the follow-up, showing no significant difference in the feeling discomfort or foreign body sensation in groin area between the two groups ( P>0.05). There was no patient occurred surgical site infection in the experiment group, and there was 1 patient in the control group occurred postoperative skin infection, which had no relationship with mesh. There was no patient in both two groups occurred fever, anaphylaxis and patch related serious adverse reaction during the follow-up. The resting visual analogue scale score, active visual analogue scale score of patients at postoperative 2 days and postoperative 18 months were 0.44±1.00, 1.28±1.46 and 0, 0 in the experiment group, versus 0.40±0.76, 1.28±1.14 and 0.24±1.20, 0.44±1.29 in the control group, respectively. There was a significant difference in the time effect of postoperative active visual analogue scale score of patients between the two groups ( Ftime=10.19, P<0.05). The thickness of the novel non-crosslinked biological mesh before implantation was 0.5?0.7 mm. Two months after operation, results of B-ultrasonic examination in groin area of 10 patients from the experiment group showed a strong echo area at the patch implant area with a thickness as 2 mm. Conclusion:Application of novel non-crosslinked biological mesh in laparoscopic inguinal hernia repair is safe and effective.

Effect and mechanism of baicalin on the proliferation and migration of human periodontal ligament stem cells induced by lipopolysaccharide / 中国药房

OBJECTIVE To explore the regulatory effects of baicalin on the proliferation and migration of human periodontal ligament stem cells （hPDLSCs） induced by lipopolysaccharide （LPS） and Janus protein tyrosine kinase 2 （JAK2）/signal transduction and transcription activator 3 （STAT3） signaling pathways. METHODS hPDLSCs were divided into control group， LPS group， different concentration baicalin groups （0.1， 1 and 10 mg/L）. ELISA method and CCK-8 assay were used to determine the contents of cell inflammatory factors [interleukin 6 （IL-6）， IL-1β and tumor necrosis factor-α （TNF-α）] and cell viability， so as to screen the optimal concentration of baicalin for follow-up pathway validation experiments. The cells were then divided into control group， LPS group， optimal baicalin concentration group and inhibitor group （10 μg/mL LPS+1 mg/L baicalin +3 μmol/L JAK2/STAT3 pathway inhibitor AG490）. After treated for 24 h， the proliferation rate of hPDLSCs， apoptosis rate， migration rate， invasion cell number， mRNA and protein expressions of Cyclin D1 and caspase-3， the expression of JAK2/STAT3 pathway-related proteins were all detected. RESULTS According to cell inflammatory factors and cell viability， 1 mg/L was selected as the optimal concentration of baicalin. Compared with control group， cell proliferation rate， migration rate， invasion cell number， Cyclin D1 mRNA and protein expression were significantly decreased in LPS group， while cell apoptosis rate， caspase-3 mRNA and protein expression， p-JAK2 and p-STAT3 protein expression were significantly increased （P＜0.05）. After treated with 1 mg/L baicalin， the above indexes were reversed significantly （P＜0.05）. The improvement of above indexes in the inhibitor group was more obvious （P＜0.05）. CONCLUSIONS Baicalin can promote the proliferation， migration and invasion of LPS-induced hPDLSCs and inhibit their apoptosis and inflammation by blocking the JAK2/STAT3 pathway.

High-throughput screening of SARS-CoV-2 main and papain-like protease inhibitors

The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for patient treatment. Here, we screened about 1.8 million small molecules against the main protease (Mpro) and papain like protease (PLpro), two major proteases in severe acute respiratory syndrome-coronavirus 2 genome, and identified 1851Mpro inhibitors and 205 PLpro inhibitors with low nmol/l activity of the best hits. Among these inhibitors, eight small molecules showed dual inhibition effects on both Mpro and PLpro, exhibiting potential as better candidates for COVID-19 treatment. The best inhibitors of each protease were tested in antiviral assay, with over 40% of Mpro inhibitors and over 20% of PLpro inhibitors showing high potency in viral inhibition with low cytotoxicity. The X-ray crystal structure of SARS-CoV-2 Mpro in complex with its potent inhibitor 4a was determined at 1.8 Å resolution. Together with docking assays, our results provide a comprehensive resource for future research on anti-SARS-CoV-2 drug development.

Establishment and Evaluation of Intestinal Injury Model of Mouse Acute Graft Versus Host Disease Based on An Organoid Technology / 中国实验血液学杂志

OBJECTIVE@#To establish an intestinal organoid model that mimic acute graft versus host disease (aGVHD) caused intestinal injuries by using aGVHD murine model serum and organoid culture system, and explore the changes of aGVHD intestine in vitro by advantage of organoid technology.@*METHODS@#20-22 g female C57BL/6 mice and 20-22 g female BALB/c mice were used as donors and recipients for bone marrow transplantation, respectively. Within 4-6 h after receiving a lethal dose (8.0 Gy) of γ ray total body irradiation, a total of 0.25 ml of murine derived bone marrow cells (1×107/mice, n=20) and spleen nucleated cells (5×106/mice, n=20) was infused to establish a mouse model of aGVHD (n=20). The aGVHD mice were anesthetized at the 7th day after transplantation, and the veinal blood was harvested by removing the eyeballs, and the serum was collected by centrifugation. The small intestinal crypts of healthy C57BL/6 mice were harvested and cultivated in 3D culture system that maintaining the growth and proliferation of intestinal stem cells in vitro. In our experiment, 5%, 10%, 20% proportions of aGVHD serum were respectively added into the organoid culture system for 3 days. The formation of small intestinal organoids were observed under an inverted microscope and the morphological characteristics of intestinal organoids in each groups were analyzed. For further evaluation, the aGVHD intestinal organoids were harvested and their pathological changes were observed. Combined with HE staining, intestinal organ morphology evaluation was performed. Combined with Alcian Blue staining, the secretion function of aGVHD intestinal organoids was observed. The distribution and changes of Lgr5+ and Clu+ intestinal stem cells in intestinal organoids were analyzed under the conditions of 5%, 10% and 20% serum concentrations by immunohistochemical stainings.@*RESULTS@#The results of HE staining showed that the integrity of intestinal organoids in the 5% concentration serum group was better than that in the 10% and 20% groups. The 5% concentration serum group showed the highest number of organoids, the highest germination rate and the lowest pathological score among experimental groups, while the 20% group exhibited severe morphological destruction and almost no germination was observed, and the pathological score was the highest among all groups(t=3.668, 4.334,5.309,P<0.05). The results of Alican blue staining showed that the secretion function of intestinal organoids in serum culture of aGVHD in the 20% group was weaker than that of the 5% group and 10% of the organoids, and there was almost no goblet cells, and mucus was stainned in the 20% aGVHD serum group. The immunohistochemical results showed that the number of Lgr5+ cells of intestinal organoids in the 5% group was more than that of the intestinal organoids in the 10% aGVHD serum group and 20% aGVHD serum group. Almost no Clu+ cells were observed in the 5% group. The Lgr5+ cells in the 20% group were seriously injuried and can not be observed. The proportion of Clu+ cells in the 20% group significantly increased.@*CONCLUSION@#The concentration of aGVHD serum in the culture system can affect the number and secretion function of intestinal organoids as well as the number of intestinal stem cells in organoids. The higher the serum concentration, the greater the risk of organoid injury, which reveal the characteristics of the formation and functional change of aGVHD intestinal organoids, and provide a novel tool for the study of intestinal injury in aGVHD.

Effect of electroacupuncture at different frequencies on brain insulin signaling transduction pathway in Alzheimer's disease mice / 中国针灸

OBJECTIVE@#To observe the effect of electroacupuncture (EA) at different frequencies on learning and memory functions, as well as the relevant proteins of brain insulin signal transduction pathway in Alzheimer's disease (AD) mice and explore the effect mechanism of EA in treatment of AD.@*METHODS@#Seventy-two SPF Kunming male mice were randomized into a blank group, a sham-operation group, a model group, a 2 Hz EA group, a 15 Hz EA group and a 30 Hz EA group, 12 mice in each one. In the model group and each EA group, AD model were established by the injection with streptozotocin (ST2) solution (8 mg/kg) into the left lateral ventricles. In the sham-operation group, 0.9% sodium chloride solution of the same volume was injected into the left lateral ventricles. After successful modeling, in each EA group, EA was applied at "Baihui" (GV 20), "Dazhui" (GV 14) and "Shenshu" (BL 23) with corresponding frequencies, once daily. One course of EA intervention consisted of 7 treatments and 2 courses were given totally at interval of 1 day. After modeling and intervention, Morris water maze test was conducted for the mice of each group. Using immunohistochemistry and Western blot method, the protein expression of insulin receptor (IR), insulin receptor substrate-1 (IRS-1) and phosphatidylinositol 3-kinase (PI3K) was detected in the hippocampal of the mice after intervention.@*RESULTS@#Compared with the blank group, in the model group, the 2 Hz, 15 Hz and 30 Hz EA groups, the escape latency and the first time of crossing the platform were all extended (P<0.01), and the number of crossing the platform was reduced (P<0.01) after modeling. When compared with the blank group, the escape latency and the first time of crossing the platform were all extended (P<0.01), and the number of crossing the platform was reduced (P<0.01) in the model group after intervention. In the 2 Hz, 15 Hz and 30 Hz EA groups, the escape latency and the first time of crossing the platform were all shortened (P<0.01), and the number of crossing the platform was increased (P<0.05, P<0.01) after intervention when compared with the model group. The escape latency and the first time of crossing the platform were all shortened (P<0.01, P<0.05), and the number of crossing the platform was increased (P<0.05) in the 15 Hz and 30 Hz EA groups in comparison with the 2 Hz EA group. The protein expression levels of IR, IRS-1 and PI3K were reduced in the model group when compared with those of the blank group (P<0.01, P<0.05); and these protein expression levels were increased in the 15 Hz and 30 Hz EA groups compared with the model group (P<0.05, P<0.01). Compared with the 2 Hz EA group, the protein expression levels of IR, IRS-1 and PI3K were all elevated in the 15 Hz and 30 Hz EA groups (P<0.05).@*CONCLUSION@#The learning and memory function of AD mice may be improved through regulating brain insulin signaling transconduction pathway with electroacupuncture, and electroacupuncture at 15 Hz and 30 Hz obtains the overall better effect compared with the intervention at 2 Hz.

Prevalence of primary biliary cholangitis in the Chinese general population and its influencing factors: A systematic review / 临床肝胆病杂志

Objective To systematically review the epidemiological studies on primary biliary cholangitis (PBC), and to investigate the prevalence rate of PBC in the Chinese general population and its influencing factors. Methods PubMed, Embase, The Cochrane Library, CNKI, and Wanfang Data were searched for articles on the epidemiology of PBC in China published up to 31th March 2022. Two researchers independently performed screening and data extraction, and then related analyses were performed. Results A total of 9 articles were included. The positive rate of AMA was 1 049.05/100 000 (ranging fr om 159.65/100 000 to 2287.40/100 000), and the prevalence rate of PBC was 123.68/100 000 (ranging from 42.70/100 000 to 276.59/100 000). The positive rate of AMA was 636.51/100 000 (ranging from 52.55/100 000 to 1 164.33/100 000) in men and 1 265.47/100 000 (ranging from 225.23/100 000 to 1 704.93/100 000) in women, with a male/female ratio of 1∶1.99 for the prevalence rate of AMA. The prevalence rate of PBC was 40.81/100 000 (ranging from 23.54/100 000 to 75.10/100 000) in men and 148.71/100 000 (ranging from 77.36/100 000 to 214.91/100 000) in women, with a male/female ratio of 1∶3.64 for the prevalence rate of PBC. Conclusion Different studies show great differences in the positive rate of AMA and the prevalence rate of PBC in the Chinese general population, which is mainly affected by sex, age, and region. The positive rate of AMA and the prevalence rate of PBC increase with age, and the patients aged ≥50 years have a significantly higher positive rate of AMA than those aged < 50 years. The positive rate of AMA is significantly higher than the prevalence rate of PBC. There are significantly more women than men in the AMA-positive population and the PBC patients, and the influence of sex on AMA is lower than that on PBC.