Liver fibrosis showed a two-phase regression rate during long-term anti-HBV therapy by three-time biopsies assessments.

BACKGROUND: Evidence has proven that liver fibrosis or even cirrhosis can be reversed by anti-HBV treatment. However, the difference of fibrosis regression rates in short-term and long-term antiviral therapy remain unclear. Therefore, we aimed to identify the dynamic changes in fibrosis regression rate in patients with three-time liver biopsies during 5 years antiviral therapy. METHODS: CHB patients with three times of liver biopsies (baseline, after 1.5-year and 5-year antiviral therapy) from a prospective cohort were enrolled. All patients were biopsy-proved Ishak stage ≥ 3 at baseline (n = 92). Fibrosis regression was defined as Ishak stage decreased ≥ 1 or predominantly regressive categorized by P-I-R score. RESULTS: Totals of 65.2% (60/92) and 80.4% (74/92) patients attained fibrosis regression after 1.5-year and 5-year therapy, respectively. Median HBV DNA level declined from 6.5 log IU/ml (baseline) to 0 log IU/ml (1.5 years and 5 years, P < 0.001). The mean level of Ishak fibrosis stage in all patients decreased from stage 4.1 (baseline) to 3.7 (1.5 years) then 3.2 (5 years). Fibrosis regression rates were 0.27 stage/year between baseline to year 1.5 and 0.14 stage/year between year 1.5 and year 5. Furthermore, for patients who attained fibrosis regression after 5-year antiviral therapy, the two-phase regression rates were 0.39 stage/year (0 year-1.5 years) and 0.20 stage/year (1.5 years-5 years). This two-phase feature of regression rate was further confirmed by fully-quantification assessment of liver fibrosis based on SHG/TPEF. CONCLUSION: During the 5 years of long-term antiviral treatment, liver fibrosis rapidly regresses in the first 1.5 years before slowing down in the following 3.5 years.

Preliminary genetic parameter estimates of meat quality traits in Hu sheep.

Because substantial numbers of Chinese consumers are prepared to pay for tender and quality lamb, meat quality traits are becoming more relevant for breeding programs for Chinese sheep breeds. The current study estimated heritabilities and genetic correlations for 13 meat quality traits recorded on lamb loins from Hu sheep. Heritability estimates ranged from 0.04 ± 0.06 for meat redness at 45 min to 0.57 ± 0.10 for drip loss, with most of the meat quality traits having moderate heritabilities. Positive genetic correlations were observed among meat color traits. Intramuscular fat (IMF) was genetically correlated with most meat quality traits, indicating that increasing IMF can favor meat pH, color, and tenderness, but would lead to increased cooking loss. Direct selection to increase IMF of loins is recommended to be included in breeding programs for Hu sheep, as it was more efficient than indirect selection on the other meat quality traits. The genetic parameters presented in this preliminary study provide valuable genetic information needed to design a breeding program aimed at improving the quality of lamb meat from Hu sheep.

Validation of the revised electronic version of RUCAM for diagnosis of DILI in Chinese patients.

BACKGROUND AIMS: The Revised Electronic Causality Assessment Method (RECAM), a computerized update of the Roussel Uclaf Causality Assessment Methodology (RUCAM), was recently proposed. In this study, we validated and compared the utility of the RECAM and RUCAM in Chinese patients with a single conventional or herbal agent-induced liver injury. METHODS: In this retrospective multicenter cohort of well-established DILI and non-DILI patients from 5 centers in China, the diagnostic performance of the RUCAM and RECAM was compared by AUC analysis. The consistency was evaluated by weighted kappa. The major causes of discrepancy were explored. RESULTS: A total of 481 DILI and 100 non-DILI patients were included. In total, 62.6% of the DILI cases were induced by conventional agents, and 37.4% were induced by herbs. The RECAM had relatively higher AUC than RUCAM for overall [0.947 (0.926-0.964) vs. 0.867 (0.836-0.893), p=0.0016], conventional agents [0.923 (0.890-0.949) vs. 0.819 (0.775-0.858), p=0.0185], and herbs [0.972 (0.941-0.989) vs.0.911 (0.866-0.944), p=0.0199]. Latency, scores associated with hepatitis B, and hepatotoxicity information of the insulting drugs were the 3 main causes for the inconsistency between RECAM and RUCAM scores. CONCLUSIONS: The RECAM had relatively better diagnostic performance than RUCAM, with a higher AUC for Chinese DILI patients. Timely updates of the LiverTox category and refinement of serum markers to exclude hepatitis B activity would further improve the applicability of RECAM in areas where the use of herbs and resolution of past HBV infections are common.

Changing spectrum and mortality disparities of etiology of liver cirrhosis in Beijing, China.

Liver cirrhosis remains a major health concern globally, but its epidemiology and etiology evolve with time. However, the changing pattern in etiology and cause of liver-related mortality for patients with cirrhosis are not fully elucidated. Herein, our aim was to characterize the temporal trend of the etiological spectrum and evaluate the impact of etiology on liver-related death among patients with compensated cirrhosis (CC) in Beijing, China. Clinical profiles of patients with CC discharged between January 2008 and December 2015 were retrieved from the Beijing hospital discharge database. The mortalities of different etiologies of cirrhosis were calculated. The risks of readmission and liver-related death associated with etiologies were evaluated by the Cox regression model. A total of 23 978 cirrhotic patients were included. The predominant cause was hepatitis B virus (HBV) (58.93%), followed by alcohol (21.35%), autoimmune (14.85%), miscellaneous etiologies (3.55%), and hepatitis C virus (HCV) (1.32%). From 2008 to 2015, the proportion of HBV-related cirrhosis decreased to 28.11%. Meanwhile, the proportions of autoimmune- and miscellaneous-related cirrhosis increased to 28.54% and 13.11%. The risk of liver-related death ranked the highest in patients with miscellaneous cirrhosis, followed by HBV-related cirrhosis, alcohol-related cirrhosis, autoimmune-related cirrhosis, and HCV-related cirrhosis. The 5-year rates of liver-related death were 22.56%, 18.99%, 18.77%, 16.01%, and 10.76%, respectively. HBV-related cirrhosis caused the highest risk of hepatocellular carcinoma (HCC)-related death, whereas alcohol- and miscellaneous-related cirrhosis caused higher risks of decompensation (DC)-related death than HBV-related cirrhosis, with hazard ratios of 1.35 (95% confidence interval [CI]: 1.24-1.48) and 1.20 (95% CI: 1.03-1.40), respectively. HBV remained a common cause of liver cirrhosis but gradually decreased. Mortality disparities existed in etiologies, with higher risks of HCC-related death in HBV-related cirrhosis, and DC-related death in alcohol- and miscellaneous-related cirrhosis.

Add-on immunosuppressive therapy may benefit selected patients with primary biliary cholangitis and autoimmune phenomena.

Background: Mildly elevated levels of transaminase and/or immunoglobulin G (IgG) are common in patients with primary biliary cholangitis (PBC). It is still unclear whether adding immunosuppressive therapy to ursodeoxycholic acid (UDCA) benefits those patients who are not fulfilling the diagnostic criteria of PBC with autoimmune hepatitis (AIH) features. Objectives: To assess the efficacy of adding immunosuppressive therapy to UDCA for patients with PBC and autoimmune phenomena but not fulfilling the diagnostic criteria of PBC with AIH features. Design: This is a retrospective-prospective cohort study in a tertiary medical center. Methods: Patients with PBC and autoimmune phenomena were defined by the elevation of IgG and/or transaminase but did not fulfill the diagnostic criteria of PBC with AIH features. We grouped these patients based on with and without add-on immunosuppressive therapy and balanced their baseline characteristics using inverse probability treatment weighting (IPTW). Results: A total of 652 patients with PBC and autoimmune phenomena were included, with a median follow-up of 4.08 years. After IPTW, the pseudo sample size in the add-on therapy and monotherapy groups was 558 and 655, respectively. After 1 year of observation, patients in the add-on therapy group had a higher biochemical response rate (normalization of transaminase and IgG levels) (49% versus 17%, p < 0.001). Furthermore, add-on therapy improved the transplant-free survival in the subgroup of patients with PBC and transaminase ⩾3 × upper limit of normal (ULN) or IgG ⩾1.3 × ULN (p = 0.033). Conclusion: Add-on immunosuppressive therapy may improve the normalization rates of transaminase and IgG levels in all patients with PBC and mildly elevated transaminase and IgG levels and the long-term outcomes in the subgroup of the patients with transaminase ⩾3 × ULN or IgG ⩾1.3 × ULN.

A look at add-on immunosuppressive therapy in primary biliary cholangitis patients Adding immunosuppressive therapy may enhance the normalization of ALT, AST and IgG levels in all PBC patients with mild elevation and improve long-term outcomes in those with more severe elevation of ALT, AST and IgG. These findings contribute to our understanding of treatment options for PBC patients with autoimmune phenomena.

Regression of Liver Fibrosis in Patients on Hepatitis B Therapy Is Associated With Decreased Liver-Related Events.

BACKGROUND & AIMS: Liver fibrosis in patients with chronic hepatitis B can regress with successful antiviral therapy. However, the long-term clinical benefits of fibrosis regression have not been fully elucidated. This study investigated the association between biopsy-proven fibrosis regression by predominantly progressive, indeterminate, and predominantly regressive (P-I-R) score and liver-related events (LREs) in chronic hepatitis B patients. METHODS: Patients with on-treatment liver biopsy and significant fibrosis/cirrhosis (Ishak stage ≥3) were included in this analysis. Fibrosis regression was evaluated according to the P-I-R score of the Beijing Classification. LREs were defined as decompensations, hepatocellular carcinoma, liver transplantation, or death. The Cox proportional hazards model was used to determine associations of fibrosis regression with LREs. RESULTS: A total of 733 patients with Ishak stages 3/4 (n = 456; 62.2%) and cirrhosis (Ishak stages 5/6; n = 277; 37.8%) by on-treatment liver biopsy were enrolled. According to the P-I-R score, fibrosis regression, indeterminate, and progression were observed in 314 (42.8%), 230 (31.4%), and 189 (25.8%) patients, respectively. The 7-year cumulative incidence of LREs was 4.1%, 8.7%, and 18.1% in regression, indeterminate, and progression, respectively (log-rank, P < .001). Compared with patients with fibrosis progression, those with fibrosis regression had a lower risk of LREs (adjusted hazard ratio, 0.40; 95% CI, 0.16-0.99; P = .047), followed by the indeterminate group (adjusted hazard ratio, 0.86; 95% CI, 0.40-1.85; P = .691). Notably, this favorable association also was observed in patients with cirrhosis or low platelet counts (<150 × 109/L). CONCLUSIONS: Antiviral therapy-induced liver fibrosis regression assessed by P-I-R score is associated with reduced LREs. This shows the utility of histologic fibrosis regression assessed by on-treatment P-I-R score as a surrogate endpoint for clinical events in patients with hepatitis B virus-related fibrosis or early cirrhosis.

Molecular characterization of SPATA6 and association of its SNPs with testicular size in sheep.

The testis is an important organ for maintaining fertility in males, and testis size is positively correlated with ejaculate volume, sperm motility, thus fertility. Spermatogenesis-associated 6 (SPATA6) is an evolutionarily conserved testis-specific gene reported in many species. However, the effect of SPATA6 expression levels on testicular development and the effect of single nucleotide polymorphisms (SNPs) on testis and epididymis phenotype in sheep have not been studied. The purpose of the research was to investigate the expression profile of SPATA6 and its effect on testicular development and to confirm the effect of SNPs on the testis and epididymis phenotype. In this study, we detected a 1245bp coding sequence (CDS) of SPATA6 and encoded 414 amino acids. The expression levels of SPATA6 were significantly higher in the testis than in other tissues and gradually increased with testis development. Moreover, the expression level in the large testis was significantly higher than that in the small testis at six months. A total of 11 SNPs were detected in the coding region of SPATA6 by cDNA-pooling sequencing and improved multiplex ligation detection reaction (iMLDR) methods. Correlation analysis showed that SNP2 (c. 3631C > G) significantly affected left epididymis weight (LEW) and right epididymis weight (REW), and SNP10 (c. 937 A > G) significantly affected REW. And the combined genotype of SNP1 (c. 4245 G > A) and SNP2 significantly affected REW. The current study concluded that SPATA6 plays an important role in testicular development and the SNPs significantly associated with the epididymis phenotype can provide molecular markers for the early selection of high-fertility Hu sheep.

Carvedilol Plus NUC for Patients With HBV-Compensated Cirrhosis Under Virological Suppression: A Randomized Open-Label Trial.

INTRODUCTION: Portal hypertension progression can be relieved after controlling the etiology of liver cirrhosis. Whether beta-blockers could additionally enhance the effects during treatment, particularly for small esophageal varices (EV), was unclear. This study aims to assess the efficacy of add-on carvedilol to delay EV progression during anti-hepatitis B virus (HBV) treatment in HBV-related cirrhosis. METHODS: This randomized controlled trial enrolled patients with virologically suppressed HBV-compensated cirrhosis and small/medium EV. The participants were randomly assigned to receive nucleos(t)ide analog (NUC) or carvedilol 12.5 mg plus NUC (1:1 allocation ratio). The primary end point was the progression rate of EV at 2 years of follow-up. RESULTS: A total of 238 patients (small EV, 77.3%) were randomized into 119 NUC and 119 carvedilol plus NUC (carvedilol [CARV] combination group). Among them, 205 patients (86.1%) completed paired endoscopies. EV progression rate was 15.5% (16/103) in the NUC group and 12.7% (13/102) in the CARV combination group (relative risk = 0.79, 95% confidence interval 0.36-1.75, P = 0.567). Subgroup analysis on medium EV showed the CARV combination group had a more favorable effect in promoting EV regression (43.5% vs 13.1%, P = 0.022) than NUC alone, but not in small cases ( P = 0.534). The incidence of liver-related events (decompensation, hepatocellular carcinoma, or death/liver transplantation) within 2 years was similar between the 2 groups (11.2% vs 10.4%, P = 0.881). DISCUSSION: The overall results did not show statistically significant differences between the added carvedilol strategy and NUC monotherapy in preventing EV progression in patients with virologically suppressed HBV-compensated cirrhosis. However, the carvedilol-added approach might offer improved outcomes specifically for patients with medium EV (NCT03736265).

Persistent steatosis correlates with decreased fibrosis regression during anti-HBV treatment in patients with chronic HBV infection.

Despite the increasing prevalence of steatosis in patients with chronic hepatitis B (CHB), whether the changes in steatosis impact fibrosis regression during antiviral therapy remain unclear. We aimed to identify the association between histological changes of steatosis and fibrosis in patients undergone antiviral treatment. Patients with paired liver biopsies before and after 78 weeks of antiviral therapy were enrolled in this study. Liver fibrosis was assessed by the Ishak score combined with Beijing Classification predominantly progressive, indeterminate, and predominately regressive score. Steatosis was evaluated by the nonalcoholic fatty liver disease activity score. Collagen in each site was quantitated by second harmonic generation/two photon excitation fluorescence technology. Serum proteomic changes after treatment were characterized by mass-based spectrometry. A total of 239 CHB patients were included and divided into four groups according to the changes in steatosis: 162 (67.8%) had no steatosis throughout, 24 (10.0%) developed new-onset steatosis, 21 (8.8%) had initial steatosis which disappeared, and 32 (13.4%) had persistent steatosis. The persistent steatosis group showed the lowest rate of fibrosis regression (14/32, 43.8%). Persistent steatosis correlated with decreased fibrosis regression significantly after adjusting for age, sex, fibrosis stage, and metabolic factors at baseline, as well as the viral response (adjusted odds ratio = 0.380, 95% confidence interval 0.145-0.996, p = 0.049). This decreased fibrosis regression was associated with accumulated collagen in the perisinusoidal area. Patients with persistent steatosis showed unique changes in glycolipid metabolism according to the serum proteomic atlas. Persistent steatosis correlated with decreased fibrosis regression during antiviral therapy in patients with CHB.

Long-course chemoradiation plus concurrent/sequential PD-1 blockade as neoadjuvant treatment for MMR-status-unscreened locally advanced rectal cancer: protocol of a multicentre, phase 2, randomised controlled trial (the POLAR-STAR trial).

INTRODUCTION: Recent preclinical studies have discovered unique synergism between radiotherapy and immune checkpoint inhibitors, which has already brought significant survival benefit in lung cancer. In locally advanced rectal cancer (LARC), neoadjuvant radiotherapy plus immune checkpoint inhibitors have also achieved surprisingly high pathological complete response (pCR) rates even in proficient mismatch-repair patients. As existing researches are all phase 2, single-cohort trials, we aim to conduct a randomised, controlled trial to further clarify the efficacy and safety of this novel combination therapy. METHODS AND ANALYSIS: Eligible patients with LARC are randomised to three arms (two experiment arms, one control arm). Patients in all arms receive long-course radiotherapy plus concurrent capecitabine as neoadjuvant therapy, as well as radical surgery. Distinguishingly, patients in arm 1 also receive anti-PD-1 (Programmed Death 1) treatment starting at Day 8 of radiation (concurrent plan), and patients in arm 2 receive anti-PD-1 treatment starting 2 weeks after completion of radiation (sequential plan). Tislelizumab (anti-PD-1) is scheduled to be administered at 200 mg each time for three consecutive times, with 3-week intervals. Randomisation is stratified by different participating centres, with a block size of 6. The primary endpoint is pCR rate, and secondary endpoints include neoadjuvant-treatment-related adverse event rate, as well as disease-free and overall survival rates at 2, 3 and 5 years postoperation. Data will be analysed with an intention-to-treat approach. ETHICS AND DISSEMINATION: This protocol has been approved by the institutional ethical committee of Beijing Friendship Hospital (the primary centre) with an identifying serial number of 2022-P2-050-01. Before publication to peer-reviewed journals, data of this research will be stored in a specially developed clinical trial database. TRIAL REGISTRATION NUMBER: NCT05245474.

Distinct on-treatment HCC risks associated with different decompensation events in HBV patients with cirrhosis.

OBJECTIVES: Long-term treatment with nucleoside analog (NA) reduces the risks for decompensation and hepatocellular carcinoma (HCC) in chronic hepatitis B (CHB) patients with compensated cirrhosis (CC). However, whether antiviral therapy has differential efficacy on the risks for decompensation and HCC is insufficiently elucidated. Therefore, we investigated the disease state transition, focusing on decompensation event-specific HCC risk in NA-treated CHB patients with CC. METHODS: We prospectively followed up on 1163 NA-treated CHB patients with CC every six months for up to seven years. The cumulative incidence and risk of HCC were analyzed by the Kaplan-Meier method and competing risk model. The multistate model was used to estimate the transition probabilities to HCC from different disease states. RESULTS: HCC predominated the first liver-related events, with a 5-year cumulative incidence of 9.0%, followed by decompensation (8.3%, including 7.9% nonbleeding decompensation and 2.4% variceal bleeding) and 0.2% death. The decompensation stage had a significantly higher 5-year cumulative HCC incidence than the CC stage (27.6% vs. 9.1%; HR = 2.42, 95% CI: 1.24, 4.71). Furthermore, nonbleeding decompensation events had a higher 5-year transition probability to HCC than bleeding (27.6% vs. 15.8%; HR = 2.69, 95% CI: 1.41, 4.17). Viral suppression modified the on-treatment transition risk to HCC (1-year: HR = 0.45, 95% CI: 0.28, 0.73; 3-year: HR = 0.23, 95% CI: 0.14, 0.38). An online calculator was developed to facilitate HCC risk stratification. CONCLUSIONS: In NA-treated CHB patients with compensated cirrhosis, the risk was higher for HCC than for decompensation; more importantly, different decompensation events conferred distinct HCC risks.

Systematic review with meta-analysis: Significant histological changes among treatment-naïve chronic hepatitis B patients with normal alanine aminotransferase levels by different criteria.

BACKGROUND: The upper limits of normal (ULNs) of ALT are not consistent across the major international guidelines which may affect the eligibility for antiviral therapy for chronic hepatitis B (CHB). AIM: To estimate the proportions of histological changes among treatment-naïve patients with CHB within differently defined ALT ULNs. METHODS: We searched PubMed and Embase up to May 15th, 2023, to identify studies of treatment-naïve CHB patients with liver biopsies. We pooled proportions of moderate to severe necroinflammation, significant fibrosis, and cirrhosis in those patients within different ALT ULNs by using random-effect models. RESULTS: We included 23 studies with 4010 participants. Within ALT ULN at 40 IU/L, the pooled proportions of moderate to severe necroinflammation, significant fibrosis, and cirrhosis were 33% (95% CI: 26%-42%), 32% (95% CI: 27%-38%), and 3% (95% CI: 1%-5%), respectively. Within ALT ULN at 30 IU/L for men and 19 IU/L for women, the pooled proportion of significant fibrosis remained at 30% (95% CI: 25%-34%; 432 participants). However, it was 21% (95% CI: 11%-37%; 361 participants) even in those within ALT ULN at 20 IU/L. Subgroup analyses suggested a significantly higher proportion of significant fibrosis among studies with prospective design or enrolled patients' mean age >35 or >40 years. CONCLUSIONS: Significant histological changes occurred in approximately 1/3 of treatment-naïve CHB patients within ALT ULN at 40 IU/L, whereas the proportion of significant fibrosis was approximately 1/5 even in those within ALT ULN at 20 IU/L.

A prospective study on the differential association of sarcopenia and frailty with health outcomes in cirrhotic patients.

BACKGROUND: To investigate the joint impact of sarcopenia and frailty on mortality and the development of decompensation in cirrhosis. METHODS: Sarcopenia was assessed using the skeletal muscle mass index (SMI) by computed tomography, whereas frailty was measured using the Fried Frailty Phenotype (FFP). Cox proportional hazard regression and competing risks analysis were used to evaluate their association with adverse outcomes. RESULTS: The prevalence of sarcopenia and frailty was 29.6% and 37.2%, respectively. Sarcopenia and frailty separately increased more than two times higher risk of all-cause mortality after adjustment for age, gender, Child-Turcotte-Pugh, and comorbidities. Co-occurrence of sarcopenia and frailty was associated with a higher incremental risk of mortality in patients with cirrhosis (HR = 4.16, 95% CI: 1.64-10.58, P = 0.003), but these two conditions didn't have significant interaction. Frailty, but not sarcopenia, was significantly associated with an increased cumulative incidence of liver-related mortality and decompensation after adjusting covariates. Subgroup analysis revealed that frailty shortened the liver-related survival of cirrhosis patients with male or higher liver severity based on MELD. CONCLUSIONS: Co-occurrence of sarcopenia and frailty increased the risk of death in cirrhosis, but these two conditions didn't have a significant interaction association. Frailty, but not sarcopenia, was associated with more adverse outcomes in cirrhotic patients.

Biomimetic polymeric transcatheter heart valve leaflets with low calcification and good regenerative ability.

Transcatheter aortic valve replacement (TAVR) technology is quickly advancing in clinic, however, as it expands to low-risk populations and younger patients (age <65 years), device durability is becoming a major challenge. Tissue-engineered heart valves (TEHVs) are a potential alternative. In this study, a bionic tri-layer tissue-engineered heart valve was constructed using poly (L-lactate-co-ε-caprolactone) (PLCL), gelatin (GEL), hyaluronic acid (HA) and silk fibroin (SF), to simulate the fibrosa, spongiosa and ventricular layer of natural heart valves. To obtain a scaffold with sufficient strength and regenerative capacity, we optimized the ratio of components of each layer. The physical and mechanical properties were tested, and the cytocompatibility, calcification deposition and regeneration potential were tested in a rat model of subcutaneous implantation. Finally, the hydrodynamic function of the new TAVR device was verified. The results demonstrated that the strength of the tri-layer valve could reach up to 10 MPa, significantly higher than that of the PLCL and mono-layer groups. Most importantly, calcification related gene expression was down-regulated in the TEHV groups compared to valvular interstitial cells (VICs) treated with calcification induced medium, and calcification levels of TEHVs in in vivo assay were below 0.5 µg mg-1. Besides, we found HA in the middle layer was very conducive to rapid cell infiltration and good angiogenesis, which ultimately promoted host tissue regeneration at 8 weeks after implantation. Collectively, we provide a bionic tri-layer electrospun leaflet with appropriate mechanical strength, low calcification and good regenerative capacity, which has great potential as a TEHV leaflet.

CfCpmd1 Regulates Pathogenicity and Sexual Development of Plus and Minus Strains in Colletotrichum fructicola Causing Glomerella Leaf Spot on Apple in China.

Colletotrichum fructicola is a devastating fungal pathogen of diverse plants. Sexually compatible plus and minus strains occur in the same ascus. However, the differentiation mechanism of plus and minus strains remains poorly understood. Here, we characterized a novel Cys2-His2-containing transcription factor CfCpmd1. The plus CfCpmd1 deletion mutant (Δ+CfCpmd1) resulted in slow hyphal growth and a fluffy cotton-like colony, and the minus deletion mutant (Δ-CfCpmd1) exhibited characters similar to the wild type (WT). Δ+CfCpmd1 led to defective perithecial formation, whereas Δ-CfCpmd1 produced more and smaller perithecia. The normal mating line was developed by pairing cultures of Δ-CfCpmd1 and plus WT, whereas a weak line was observed between Δ+CfCpmd1 and minus WT. Conidial production was completely abolished in both plus and minus mutants. When inoculated on non-wounded apple leaves with mycelial plugs, Δ-CfCpmd1 was nonpathogenic because of failure to develop conidia and appressoria, while Δ+CfCpmd1 could infect apple leaves by appressoria differentiated directly from hyphal tips, even though no conidia formed. Collectively, our results demonstrate that CfCpmd1 of C. fructicola is an important gene related to plus and minus strain differentiation, which also affects hyphal growth, sporulation, appressorium formation, and pathogenicity.

Two Putative Pheromone Receptors, but Not Their Cognate Pheromones, Regulate Female Fertility in the Atypical Mating Fungus Colletotrichum fructicola.

Colletotrichum fungi are a group of damaging phytopathogens with atypical mating type loci (harboring only MAT1-2-1 but not MAT1-1-1) and complex sexual behaviors. Sex pheromones and their cognate G-protein-coupled receptors are conserved regulators of fungal mating. These genes, however, lose function frequently among Colletotrichum species, indicating a possibility that pheromone signaling is dispensable for Colletotrichum sexual reproduction. We have identified two putative pheromone-receptor pairs (PPG1:PRE2, PPG2:PRE1) in C. fructicola, a species that exhibits plus-to-minus mating type switching and plus-minus-mediated mating line development. Here, we report the generation and characterization of gene-deletion mutants for all four genes in both plus and minus strain backgrounds. Single-gene deletion of pre1 or pre2 had no effect on sexual development, whereas their double deletion caused self-sterility in both the plus and minus strains. Moreover, double deletion of pre1 and pre2 caused female sterility in plus-minus outcrossing. Double deletion of pre1 and pre2, however, did not inhibit perithecial differentiation or plus-minus-mediated enhancement of perithecial differentiation. Contrary to the results with pre1 and pre2, double deletion of ppg1 and ppg2 had no effect on sexual compatibility, development, or fecundity. We concluded that pre1 and pre2 coordinately regulate C. fructicola mating by recognizing novel signal molecule(s) distinct from canonical Ascomycota pheromones. The contrasting importance between pheromone receptors and their cognate pheromones highlights the complicated nature of sex regulation in Colletotrichum fungi.

Hepatocellular carcinoma prediction model performance decreases with long-term antiviral therapy in chronic hepatitis B patients.

BACKGROUND/AIMS: Existing hepatocellular carcinoma (HCC) prediction models are derived mainly from pretreatment or early on-treatment parameters. We reassessed the dynamic changes in the performance of 17 HCC models in patients with chronic hepatitis B (CHB) during long-term antiviral therapy (AVT). METHODS: Among 987 CHB patients administered long-term entecavir therapy, 660 patients had 8 years of follow-up data. Model scores were calculated using on-treatment values at 2.5, 3, 3.5, 4, 4.5, and 5 years of AVT to predict threeyear HCC occurrence. Model performance was assessed with the area under the receiver operating curve (AUROC). The original model cutoffs to distinguish different levels of HCC risk were evaluated by the log-rank test. RESULTS: The AUROCs of the 17 HCC models varied from 0.51 to 0.78 when using on-treatment scores from years 2.5 to 5. Models with a cirrhosis variable showed numerically higher AUROCs (pooled at 0.65-0.73 for treated, untreated, or mixed treatment models) than models without (treated or mixed models: 0.61-0.68; untreated models: 0.51-0.59). Stratification into low, intermediate, and high-risk levels using the original cutoff values could no longer reflect the true HCC incidence using scores after 3.5 years of AVT for models without cirrhosis and after 4 years of AVT for models with cirrhosis. CONCLUSION: The performance of existing HCC prediction models, especially models without the cirrhosis variable, decreased in CHB patients on long-term AVT. The optimization of existing models or the development of novel models for better HCC prediction during long-term AVT is warranted.

A Thermostable Type I Collagen from Swim Bladder of Silver Carp (Hypophthalmichthys molitrix).

As a major component of the extracellular matrix, collagen has been used as a biomaterial for many purposes including tissue engineering. Commercial collagen derived from mammals is associated with a risk of prion diseases and religious restrictions, while fish-derived collagen can avoid such issues. In addition, fish-derived collagen is widely available and low-cost; however, it often suffers from poor thermal stability, which limits its biomedical application. In this study, collagen with a high thermal stability was successfully extracted from the swim bladder of silver carp (Hypophthalmichthys molitrix) (SCC). The results demonstrated that it was a type I collagen with high purity and well-preserved triple-helix structure. Amino acid composition assay showed that the amounts of threonine, methionine, isoleucine and phenylalanine in the collagen of swim bladder of silver carp were higher than those of bovine pericardium. After adding salt solution, swim-bladder-derived collagen could form fine and dense collagen fibers. In particular, SCC exhibited a higher thermal denaturation temperature (40.08 °C) compared with collagens from the swim bladder of grass carp (Ctenopharyngodon idellus) (GCC, 34.40 °C), bovine pericardium (BPC, 34.47 °C) and mouse tail (MTC, 37.11 °C). Furthermore, SCC also showed DPPH radical scavenging ability and reducing power. These results indicate that SCC presents a promising alternative source of mammalian collagen for pharmaceutical and biomedical applications.

Impact of polyethylene glycol loxenatide on cardiovascular outcomes in patients with type 2 diabetes: study protocol for a multicentre, randomised, double-blind, placebo-controlled trial (BALANCE-3).

INTRODUCTION: Recent cardiovascular outcomes trials have demonstrated that glucagon-like peptide 1 receptor agonist (GLP-1RA) decreases the incidence of major adverse cardiovascular events (MACEs) in individuals with type 2 diabetes mellitus (T2DM). Polyethylene glycol loxenatide (PEG-Loxe) is a once-weekly GLP-1RA obtained by modifying exendin-4. No clinical trials have been designed to assess the impact of PEG-Loxe on cardiovascular (CV) outcomes in individuals with T2DM. This trial aims to test the hypothesis that compared with placebo, PEG-Loxe treatment does not result in an unacceptable increase in CV risk in individuals with T2DM. METHODS AND ANALYSIS: This study is a multicentre, randomised, double-blind, placebo-controlled trial. Patients with T2DM who fulfilled the inclusion criteria were randomly divided to receive weekly administration of either PEG-Loxe 0.2 mg or placebo (1:1 ratio). The randomisation was stratified according to utilisation of sodium-glucose cotransporter 2 inhibitors, history of CV disease and body mass index. The research period is expected to be 3 years, with a 1-year recruitment period and a 2-year follow-up period. The primary outcome is the occurrence of the first MACE, described as CV death, non-fatal myocardial infarction or non-fatal stroke. The statistical analyses were undertaken on the intent-to-treat patient. The primary outcome was evaluated using a Cox proportional hazards model with treatment and randomisation strata as the covariates. ETHICS AND DISSEMINATION: The current research has been authorised by the Ethics Committee of Tianjin Medical University Chu Hsien-I Memorial Hospital (approval number: ZXYJNYYhMEC2022-2). Researchers must acquire informed consent from every participant before conducting any protocol-associated procedures. The findings of this study will be published in a peer-reviewed journal. TRIAL REGISTRATION NUMBER: ChiCTR2200056410.