Development and application of one-step multiplex Real-Time PCR for detection of three main pathogens associated with bovine neonatal diarrhea.

Introduction: Neonatal calf diarrhea (NCD) is one of the most common diseases in calves, causing huge economic and productivity losses to the bovine industry worldwide. The main pathogens include bovine rotavirus (BRV), bovine coronavirus (BCoV), and Enterotoxigenic Escherichia coli (ETEC) K99. Since multiple infectious agents can be involved in calf diarrhea, detecting each causative agent by traditional methods is laborious and expensive. Methods: In this study, we developed a one-step multiplex Real-Time PCR assay to simultaneously detect BRV, BCoV, and E. coli K99+. The assay performance on field samples was evaluated on 1100 rectal swabs of diseased cattle with diarrhea symptoms and compared with the conventional gel-based RT-PCR assay detect BRV, BCoV, and E. coli K99+. Results: The established assay could specifically detect the target pathogens without cross-reactivity with other pathogens. A single real-time PCR can detect ~1 copy/µL for each pathogen, and multiplex real-time PCR has a detection limit of 10 copies/µL. Reproducibility as measured by standard deviation and coefficient of variation were desirable. The triple real-time PCR method established in this study was compared with gel-based PT-PCR. Both methods are reasonably consistent, while the real-time PCR assay was more sensitive and could rapidly distinguish these three pathogens in one tube. Analysis of surveillance data showed that BRV and BCoV are major enteric viral pathogens accounting for calves' diarrhea in China. Discussion: The established assay has excellent specificity and sensitivity and was suitable for clinical application. The robustness and high-throughput performance of the developed assay make it a powerful tool in diagnostic applications and calf diarrhea research. ​.

Elevated ALOX12 in renal tissue predicts progression in diabetic kidney disease.

Diabetic kidney disease (DKD) is one of the major causes of end-stage renal disease and one of the significant complications of diabetes. This study aims to identify the main differentially expressed genes in DKD from transcriptome sequencing results and analyze their diagnostic value. The present study sequenced db/m mouse and db/db mouse to determine the ALOX12 genetic changes related to DKD. After preliminary validation, ALOX12 levels were significantly elevated in the blood of DKD patients, but not during disease progression. Moreover, urine ALOX12 was increased only in macroalbuminuria patients. Therefore, to visualize the diagnostic efficacy of ALOX12 on the onset and progression of renal injury in DKD, we collected kidney tissue from patients for immunohistochemical staining. ALOX12 was increased in the kidneys of patients with DKD and was more elevated in macroalbuminuria patients. Clinical chemical and pathological data analysis indicated a correlation between ALOX12 protein expression and renal tubule injury. Further immunofluorescence double staining showed that ALOX12 was expressed in both proximal tubules and distal tubules. Finally, the diagnostic value of the identified gene in the progression of DKD was assessed using receiver operating characteristic (ROC) curve analysis. The area under the curve (AUC) value for ALOX12 in the diagnosis of DKD entering the macroalbuminuria stage was 0.736, suggesting that ALOX12 has good diagnostic efficacy. During the development of DKD, the expression levels of ALOX12 in renal tubules were significantly increased and can be used as one of the predictors of the progression to macroalbuminuria in patients with DKD.

H10Nx avian influenza viruses detected in wild birds in China pose potential threat to mammals.

H10 subtype avian influenza viruses (AIVs) have been isolated from wild and domestic avian species worldwide and have occasionally crossed the species barrier to mammalian hosts. Fatal human cases of H10N8 infections and the recent detection of human H10N3 infections have drawn widespread public attention. In this study, 25 H10Nx viruses were isolated from wild waterfowl in China during a long-term surveillance of AIVs. We conducted phylogenetic and phylogeographic studies of the hemagglutinin (HA) genes of global H10 viruses to determine the spatiotemporal patterns of spread and the roles of different hosts in viral transmission. We found the pattern of AIV transmission from wild birds to poultry to humans, and Anatidae have acted as the seeding population in the spread of the virus. Phylogenetic incongruence indicated complex reassortment events and our isolates were divided into eight genotypes (G1-8). We also found that the HA genes of the G8 viruses belonged to the North American lineage, indicating that intercontinental gene flow has occurred. Their receptor-binding specificity showed that the G1/4/5/6/7/8 viruses bind to both human-type α2,6-linked sialic acid receptors and avian-type α2,3-linked sialic acid receptors. Mouse studies indicated that the H10Nx isolates replicated efficiently in the respiratory system without preadaptation, but showed low pathogenicity in mice. The H10Nx isolates showed no (G2/4/7) or low pathogenicity (G1/3/5/6/8) in chickens, and the G6 and G8 viruses could be transmitted to chickens through direct contact. The asymptomatic shedding of these wild-bird-origin H10Nx isolates in chickens and their good adaptation in mice should increase the ease of their transmission to humans, and they therefore pose a threat to public health. Our findings demonstrate a further understanding of wild bird-origin H10 viruses and provide information for the continuous surveillance of H10 subtype viruses.

Clinical significance of Interleukin 17 receptor E in diabetic nephropathy.

OBJECTIVE: Diabetic nephropathy (DN) is a leading cause of end-stage renal disease. Since there are limited therapeutic options available for the prevention of DN progression, it is imperative to explore novel differentially expressed genes and therapeutic targets for DN. METHODS: In this study, mice kidney tissue were subjected to transcriptome sequencing and the results were analysed using bioinformatics methods. Interleukin 17 receptor E (IL-17RE) was screened from the sequencing data and its expression was validated in the animal tissues and a cross-sectional clinical study. Fifty-five DN patients were enrolled and further subdivided into two groups based on the urinary albumin-to-creatinine ratio (UACR). Two control groups were used for comparison (minimal change disease group, 12 patients; normal control group, 6 patients). Correlation analysis was conducted to study the relationship between IL-17RE expression and the clinicopathological indices. Logistic regression and receiver operating characteristic (ROC) curve analyses were conducted to evaluate the diagnostic value. RESULTS: IL-17RE expression was significantly higher in db/db mice and the kidney tissues of DN patients than the control group. IL-17RE protein levels in the kidney tissues were strongly correlated with neutrophil gelatinase-associated lipocalin (NGAL) levels, UACR, and certain clinicopathological indices. IL-17RE levels, total cholesterol (TC) levels, and glomerular lesions were independent risk factors for macroalbuminuria. ROC curves showed a good detection value for IL-17RE in macroalbuminuria (area under the curve = 0.861). CONCLUSION: The results of this study provide novel insights into DN pathogenesis. Kidney IL-17RE expression levels were associated with DN disease severity and albuminuria.

17ß-hydroxysteroid dehydrogenases in the progression of nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) has become a worldwide epidemic and a major public health problem, with a prevalence of approximately 25%. The pathogenesis of NAFLD is complex and may be affected by the environment and susceptible genetic factors, resulting in a highly variable disease course and no approved drugs in the clinic. Notably, 17ß-hydroxysteroid dehydrogenase type 13 (HSD17B13), which belongs to the 17ß-hydroxysteroid dehydrogenase superfamily (HSD17Bs), is closely related to the clinical outcome of liver disease. HSD17Bs consists of fifteen members, most related to steroid and lipid metabolism, and may have the same biological function as HSD17B13. In this review, we highlight recent advances in basic research on the functional activities, major substrates, and key roles of HSD17Bs in the progression of NAFLD to develop innovative anti-NAFLD drugs targeting HSD17Bs.

Koopman-Based MPC With Learned Dynamics: Hierarchical Neural Network Approach.

This article presents a data-driven control strategy for nonlinear dynamical systems, enabling the construction of a Koopman-based linear system associated with nonlinear dynamics. The primary idea is to apply the deep learning technique to the Koopman framework for globally linearizing nonlinear dynamics and impose a Koopman-based model predictive control (MPC) approach to stabilize the nonlinear dynamical systems. In this work, we first generalize the Koopman framework to nonlinear control systems, enabling comprehensive linear analysis and control methods to be effective for nonlinear systems. We next present a hierarchical neural network (HNN) approach to deal with the crucial challenge of the finite-dimensional Koopman representation approximation. In particular, a scale-invariant constrained network in the HNN includes four modules, in which a predictor module and a linear module can accurately approximate the finite Koopman eigenfunctions and Koopman operator, respectively, thus forming the lifted linear system. Then, we design the Koopman-based MPC scheme for controlling nonlinear systems with constraints by adopting the modified MPC with a saturation-like function on the lifted linear system. Importantly, the Koopman-based MPC enjoys higher computational efficiency compared to the classical linear MPC and nonlinear MPC methods. Finally, a physical experiment on an overhead crane system is provided to demonstrate the effectiveness of the proposed data-driven control framework.

Multifaceted roles of zinc finger proteins in regulating various agronomic traits in rice.

Rice is an important cereal crop, which provides staple food for more than half of the world's population. To meet the demand of the ever-growing population in the next few decades, an extra increase in rice yield is an urgent need. Given that various agronomic traits contribute to the yield of rice, deciphering the key regulators involved in multiple agronomic trait formation is particularly important. As a superfamily of transcription factors, zinc finger proteins participate in regulating multiple genes in almost every stage of rice growth and development. Therefore, understanding zinc finger proteins underlying regulatory network would provide insights into the regulation of agronomic traits in rice. To this end, we intend to summarize the current advances in zinc finger proteins, with emphasis on C2H2 and CCCH proteins, and then discuss their potential in improving rice yield.

Emodin-induced hepatotoxicity is enhanced by 3-methylcholanthrene through activating aryl hydrocarbon receptor and inducing CYP1A1 in vitro and in vivo.

BACKGROUND & AIMS: Polygonum multiflorum Thunb. (PMT) is the most common traditional Chinese medicine used to treat multiple diseases, and the hepatotoxicity caused by PMT has made great concern around world. Recent results showed that emodin is the potential toxic components of PMT, but the molecular mechanisms of emodin on liver toxicity remain to be elucidated. METHODS: Evaluation of parent- and metabolite-induced cytotoxicity in emodin were compared in L02 cells and mouse model from the perspective of drug metabolizing enzymes. The effect and mechanism of emodin-induced hepatotoxicity were analyzed using electrophoretic mobility shift, promoter reporter, and high content screening. RESULTS: We showed that emodin treatment (360 mg/kg in mice, 50 µM in L02 cells) induced hepatotoxicity and enhanced reactive oxidative stress (ROS) level. Importantly, emodin-induced ROS accumulation and hepatotoxicity were attenuated in the condition of CH223191, a selective inhibitor of aryl hydrocarbon receptor (AhR), and aggravated by 3-methylcholanthrene, a selective activator of AhR. Interestingly, we performed the study on ROS mediated ER stress and mitochondrial dysfunction in emodin-induced hepatotoxicity, the results showed that emodin can decrease MMP and trigger ER stress with Ca2+ overloading and the expression of ATF4 increasing, further resulted with increased apoptosis in L02 cells and mice mortality rate, while the changes were alleviated by CH223191. Furthermore, the 5-hydroxyemodin, a metabolite by emodin through CYP1A2 enzyme, showed more severe hepatotoxicity compared to emodin. CONCLUSIONS: Our results validated that the metabolism of emodin to 5-hydroxyemodin by CYP1A played an important role in the hepatocellular toxicity of emodin and provided evidence that CYP1A1 and AhR could be used to predict and validate patient-specific liver injury of PMT or other herbs containing emodin.

Bicyclol ameliorates advanced liver diseases in murine models via inhibiting the IL-6/STAT3 signaling pathway.

Bicyclol, a synthetic hepatoprotective and anti-inflammatory agent approved in China, was widely used to treat various hepatitis accompanied by elevated serum aminotransferases. However, the pharmacological effects and mechanisms of bicyclol on advanced liver diseases, such as fibrosis/cirrhosis and hepatocellular carcinoma (HCC), remain to be explored. Here, we revealed that bicyclol prevents from formatting severe fibrosis, slows the progression of moderate liver fibrosis, accelerates the regression of moderate liver fibrosis, decreases the malignancy of HCC in rat models induced by diethylnitrosamine (DEN), and also blocks steatohepatitis to HCC in mice induced by western diet plus carbon tetrachloride and DEN. The detailed pharmacological mechanism showed that bicyclol alleviates chronic progressive liver diseases by inhibiting the levels of IL-6 and subsequent phosphorylated STAT3. Conclusion: Bicyclol plays significant protective roles in multiply stages of fibrosis/cirrhosis-HCC and nonalcoholic fatty liver disease-related HCC via inhibiting IL-6/STAT3 signaling pathway. Therefore, bicyclol might be a promising therapeutic strategy for treating advanced liver diseases.

Down-Regulating the High Level of 17-Beta-Hydroxysteroid Dehydrogenase 13 Plays a Therapeutic Role for Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide, and there is no specific drug to treat it. Recent results showed that 17-beta-hydroxysteroid dehydrogenase type 13 (HSD17B13) is associated with liver diseases, but these conclusions are controversial. Here, we showed that HSD17B13 was more highly expressed in the livers of NAFLD patients, and high expression was induced in the livers of murine NAFLD models and cultural hepatocytes treated using various etiologies. The high HSD17B13 expression in the hepatocytes facilitated the progression of NAFLD by directly stabilizing the intracellular lipid drops and by indirectly activating hepatic stellate cells. When HSD17B13 was overexpressed in the liver, it aggravated liver steatosis and fibrosis in mice fed with a high-fat diet, while down-regulated the high expression of HSD17B13 by short hairpin RNAs produced a therapeutic effect in the NAFLD mice. We concluded that high HSD17B13 expression is a good target for the development of drugs to treat NAFLD.

Highly Pathogenic Avian Influenza A(H5N8) Clade 2.3.4.4b Viruses in Satellite-Tracked Wild Ducks, Ningxia, China, 2020.

During October 2020, we identified 13 highly pathogenic avian influenza A(H5N8) clade 2.3.4.4b viruses from wild ducks in Ningxia, China. These viruses were genetically related to H5N8 viruses circulating mainly in poultry in Europe during early 2020. We also determined movements of H5N8 virus‒infected wild ducks and evidence for spreading of viruses.

Highly Pathogenic Avian Influenza A(H5Nx) Virus of Clade 2.3.4.4b Emerging in Tibet, China, 2021.

H5N8 and H5N1 highly pathogenic avian influenza viruses (AIVs) of clade 2.3.4.4b were isolated from dead migratory birds and fecal samples collected in Tibet, China, in May 2021. Phylogenetic analyses showed that the viruses isolated in this study may have spread from wintering or stopover grounds of migratory birds in South Asia. We monitored two disparate clade 2.3.4.4b H5Nx viruses in migratory birds in Tibet during their breeding season. The data revealed that breeding grounds may exhibit a potential pooling effect among avian influenza viruses in different migratory populations. IMPORTANCE In this study, 15 H5N8 and two H5N1 highly pathogenic avian influenza viruses of clade 2.3.4.4b were isolated from dead migratory birds and fecal samples in Tibet, China. Isolates of H5N1 virus of clade 2.3.4.4b have been rarely reported in China. Our findings highlight that breeding grounds may exhibit a potential pooling effect among avian influenza viruses (AIVs) in different migratory populations. In addition to intensification of the surveillance of AIVs in migratory birds in Tibet, China, international cooperation should be strengthened.

Anaphylactic Rare Saponins Separated from Panax notoginseng Saponin and a Proteomic Approach to Their Anaphylactic Mechanism.

In recent years, many traditional Chinese medicine injections based on Panax notoginseng saponin (PNS) have been reported to cause anaphylaxis. Previous studies on the anaphylactic saponins of PNS and their mechanism are inadequate. In this study, potential anaphylactic saponins were obtained by the separation of PNS and preparation of each individual component through comprehensive techniques, such as liquid chromatography, preparative chromatography, HPLC, NMR, and MS. The anaphylactic abilities of these saponins were tested using RBL-2H3 cells via a ß-hexosaminidase release rate test. The results for the mechanism of anaphylaxis were obtained by a proteomic analysis using RBL-2H3 cells. The results indicate that, among all the saponins prepared, gypenoside LXXV and notoginsenoside T5 showed strong anaphylactic abilities and notoginsenoside ST-4 and ginsenoside Rk3 showed weak anaphylactic abilities. These 4 saponins can induce anaphylaxis via direct stimulation of effector cells. The gene oncology enrichment analysis results showed that, among these saponins, only gypenoside LXXV was related to organelles of the endoplasmic reticulum and Golgi apparatus and biological processes in response to organic cyclic compounds. Four proteins in RBL-2H3 cells with the accession numbers A0A0G2JWQ0, D3ZL85, D4A5G8, and Q8K3F0 were identified as crucial proteins in the anaphylactic process. This research will help traditional Chinese medicine injection manufacturers strengthen their quality control and ensure the safety of anaphylactic saponins.

Combined Use of Bicyclol and Berberine Alleviates Mouse Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD), ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), is a liver disease worldwide without approved therapeutic drugs. Anti-inflammatory and hepatoprotective drug bicyclol and multi-pharmacological active drug berberine, respectively, have shown beneficial effects on NAFLD in murine nutritional models and patients, though the therapeutic mechanisms remain to be illustrated. Here, we investigated the combined effects of bicyclol and berberine on mouse steatosis induced by Western diet (WD), and NASH induced by WD/CCl4. The combined use of these was rather safe and better reduced the levels of transaminase in serum and triglycerides and cholesterol in the liver than their respective monotherapy, accompanied with more significantly attenuating hepatic inflammation, steatosis, and ballooning in mice with steatosis and NASH. The combined therapy also significantly inhibited fibrogenesis, characterized by the decreased hepatic collagen deposition and fibrotic surface. As per mechanism, bicyclol enhanced lipolysis and ß-oxidation through restoring the p62-Nrf2-CES2 signaling axis and p62-Nrf2-PPARα signaling axis, respectively, while berberine suppressed de novo lipogenesis through downregulating the expression of acetyl-CoA carboxylase and fatty acid synthetase, along with enrichment of lipid metabolism-related Bacteroidaceae (family) and Bacteroides (genus). Of note, the combined use of bicyclol and berberine did not influence each other but enhanced the overall therapeutic role in the amelioration of NAFLD. Conclusion: Combined use of bicyclol and berberine might be a new available strategy to treat NAFLD.

Emergence, prevalence, and evolution of H5N8 avian influenza viruses in central China, 2020.

Highly pathogenic influenza A(H5N8) viruses have caused several worldwide outbreaks in birds and are able cross the species barrier to infect humans, posing a substantial threat to public health. After the first detection of H5N8 viruses in deceased swans in Inner Mongolia, we performed early warning and active monitoring along swan migration routes in central China. We isolated and sequenced 42 avian influenza viruses, including 40 H5N8 viruses, 1 H5N2 virus, and 1 H9N2 virus, in central China. Our H5N8 viruses isolated in swan stopover sites and wintering grounds showed high nucleotide homologies in the whole genome, revealing a common evolutionary source. Phylogenetic analysis revealed that the H5 viruses of clade 2.3.4.4b prevalent in 2020 have further diverged into two sub-clades: b1 and b2. The phylogeographic analysis also showed that the viruses of sub-clade b2 most likely originated from poultry in Russia. Notably, whooper swans were found to be responsible for the introduction of sub-clade b2 viruses in central China; whooper and tundra swans play a role in viral spread in the Yellow River Basin and the Yangtze River Basin, respectively. Our findings highlight swans as an indicator species for transborder spreading and monitoring of the H5N8 virus.

TGF-ß isoforms inhibit hepatitis C virus propagation in transforming growth factor beta/SMAD protein signalling pathway dependent and independent manners.

Transforming growth factor beta (TGF-ß) plays an important role in the viral liver disease progression via controlling viral propagation and mediating inflammation-associated responses. However, the antiviral activities and mechanisms of TGF-ß isoforms, including TGF-ß1, TGF-ß2 and TGF-ß3, remain unclear. Here, we demonstrated that all of the three TGF-ß isoforms were increased in Huh7.5 cells infected by hepatitis C virus (HCV), but in turn, the elevated TGF-ß isoforms could inhibit HCV propagation with different potency in infectious HCV cell culture system. TGF-ß isoforms suppressed HCV propagation through interrupting several different stages in the whole HCV life cycle, including virus entry and intracellular replication, in TGF-ß/SMAD signalling pathway-dependent and TGF-ß/SMAD signalling pathway-independent manners. TGF-ß isoforms showed additional anti-HCV activities when combined with each other. However, the elevated TGF-ß1 and TGF-ß2, not TGF-ß3, could also induce liver fibrosis with a high expression of type I collagen alpha-1 and α-smooth muscle actin in LX-2 cells. Our results showed a new insight into TGF-ß isoforms in the HCV-related liver disease progression.

Combined identification of ARID1A, CSMD1, and SENP3 as effective prognostic biomarkers for hepatocellular carcinoma.

BACKGROUND: The current study aimed to understand the genetic landscape and investigate the diagnostic and prognostic biomarkers of primary hepatocellular carcinoma (HCC). METHODS: A cohort of 36 Chinese HCC samples with hepatitis B virus (HBV) infection was examined by whole-exome sequencing (WES). Prognosis-related alterations were identified and further verified in the TCGA database and GSE65372 profiles in the GEO database. A Chinese replication cohort of 180 HCC samples with HBV infection was collected to evaluate the candidate genes by immunohistochemical analysis. A receiver operating characteristic (ROC) curve analysis evaluated the prognostic power of candidate genes. Finally, EdU and transwell invasion assay were performed to detect the function of candidate genes. RESULTS: A total of 11 novel genes showed a significant association with HCC in the discovery cohort. The data were verified using the GEO and TCGA databases, and the expression of ARID1A, CSMD1, and SENP was evaluated in the replication cohort. Furthermore, ARID1A, CSMD1, and SENP3 are effective prognostic biomarkers for HCC patients in the replication population. CONCLUSIONS: Molecular heterogeneity was detected in HCC patients, and ARID1A, CSMD1, and SENP3 were identified as effective HCC prognosis biomarkers. CSMD1 prevents HCC by suppressing cell invasion.

MRWMDA: A novel framework to infer miRNA-disease associations.

MicroRNAs (miRNAs) are widely involved in a series of significant biological processes, which have been revealed and verified by accumulating experimental studies. The computational inference of the correlation between miRNAs and diseases is essential to facilitate the detection of disease biomarkers for disease diagnosis, prevention, treatment and prognosis. In this paper, a model with Multiple use of Random Walk with restart algorithm was introduced for the prediction of the MiRNA-Disease Association (MRWMDA). Based on diverse similarity measures, the model first implemented the random walk with restart (RWR) algorithm on the integrated similarity network to construct the topological similarity of miRNAs and diseases, which took full advantage of the network topology information. Then, the RWR algorithm was applied in the miRNA topological similarity network, and a steady probability of each miRNA-disease pair was obtained to prioritize miRNA candidates. In particular, the initial probability of the RWR algorithm was determined by utilizing the combination of the recommendation algorithm and the maximum similarity method. The proposed model achieved significant improvement in prediction compared with previous models, with an AUC of 0.9353 and an AUPR of 0.4809. In addition, case studies of breast neoplasms and lung neoplasms representing different disease types further demonstrated the excellent ability of MRWMDA in detecting potential disease-associated miRNAs. These performance analyses indicated that MRWMDA could be an effective and powerful biological computational tool in relevant biomedical studies.

Role of selected criteria and preventive chemotherapy in tumor recurrence after liver transplantation.

BACKGROUND: Long-term survival after liver transplantation (LT) for hepatocellular carcinoma (HCC) patients remains poor because of tumor recurrence. To improve the prognosis of HCC patients after LT, we aimed to identify different transplantation criteria and risk factors related to tumor recurrence and evaluate the effect of preventive chemotherapy in a single center. METHODS: In total, data on 20 variables and the survival of 199 patients with primary HCC who underwent LT between 2005 and 2015 were included for analysis. The patients were divided into the following three groups: Group 1, within the Milan and Hangzhou criteria (n = 51); Group 2, beyond the Milan but within the Hangzhou criteria (n = 36); and Group 3, beyond the Milan and Hangzhou criteria (n = 112). Survival probabilities for the three groups were calculated using multivariate Cox regression analysis. The association between preventive therapy and HCC-recurrence after LT was analyzed by multiple logistic regression analysis. RESULTS: Child-Pugh stage C and hepatitis B virus (HBV) infection were independent risk factors for patients with tumor recurrence who did not meet the Milan criteria. The overall survival rates of the 199 patients showed statistically significant differences among the three groups (P < 0.001). Moreover, no significant difference was noted in the survival rate between Group 1 and Group 2 (P > 0.05). Multivariate logistic regression analysis showed that postoperative prophylactic chemotherapy reduced the risk of tumor recurrence in patients who did not meet the Hangzhou and Milan criteria (OR = 0.478; 95% CI: 0.308-0.741; P = 0.001). CONCLUSIONS: Child-Pugh classification and HBV infection were the independent risk factors of tumor recurrence in HCC patients with LT. The Hangzhou criteria were effective and analogous compared with the Milan criteria. Preventive chemotherapy significantly reduced the risk of recurrence and prolonged the survival time for HCC patients beyond the Milan and Hangzhou criteria after LT.

Attenuating ischemia/reperfusion injury in rat cardiac transplantation by intracoronary infusion with siRNA cocktail solution.

Tumor necrosis factor-α (TNF-α), caspase-8, and complement component 5a receptor (C5aR) are known to play a crucial role in the myocardial ischemia/reperfusion (I/R) injury in cardiac transplantation. We hypothesized that the intracoronary infusion of TNF-α, caspase-8, and C5aR small interfering RNAs (siRNA) would protect cardiac allograft function and improve graft survival from I/R injury-induced organ failure. I/R injury of cardiac allograft was induced by syngeneic rat cardiac transplantation, in which the transplanted hearts were infused with saline or different amounts of siRNA cocktail solution targeting TNF-α, caspase-8, and C5aR via coronary arteries, and subsequently subjected to 18 h of preservation at 4°C in histidine-tryptophan-ketoglutarate (HTK) solution. The effects of siRNA cocktail solution on prolonged cold I/R injury were determined by assessing graft survival, histopathological changes, myeloperoxidase (MPO) activity, and malondialdehyde (MDA) concentration. The perfused siRNA cocktail solution successfully knocked down the expression of TNF-α, caspase-8, and C5aR in vitro and in vivo. Approximately 91.7% of control hearts that underwent 18 h of cold ischemia ceased their function after transplantation; however, 87.5% of cardiac allografts from the highest dose siRNA cocktail solution-pretreated hearts survived >14 days and exhibited minimal histological changes, with minimal cellular infiltration, interstitial edema, and inflammation and maximal reduced MPO activity and MDA concentration in the cardiac allograft. We demonstrated the feasibility and efficiency of infusion of TNF-α, caspase-8, and C5aR siRNA via the intracoronary route as a promising strategy for gene silencing against I/R injury in cardiac transplantation.