A pilot study on glutamate receptor and carrier gene variants and risk of childhood autism spectrum.

Imbalanced glutamate signaling has been implicated in the development of autism spectrum disorder (ASD). This case-control study was to examine single nucleotide polymorphisms (SNPs) in glutamate receptor and carrier genes and determine their association with childhood ASD in a Chinese Han population. A total of 12 SNPs in genes encoding glutamate receptors (GRM7 and GRM8) and carriers (SLC1A1 and SLC25A12) were examined in 249 autistic children and 353 healthy controls. The Childhood Autism Rating Scale (CARS) and its verbal communication domain were applied to evaluate the severity of the disease and language impairment, respectively. The T allele of rs2292813 in the SLC25A12 gene was significantly associated with an increased risk of ASD (odds ratio (OD) = 1.7, 95% confidence interval (CI): 1.1-2.6, P = 0.0107). Neither the genotypes nor allele distributions of other SNPs were associated with the risk of ASD. Notably, rs1800656 and rs2237731 in the GRM8 gene, but not other SNPs, were related to the severity of language impairment. All SNPs were not correlated with the overall severity of ASD. Our findings support associations between the SLC25A12 gene variant and the risk of childhood ASD, and between the GRM8 gene variant and the severity of language impairment in the Chinese Han population.

Treatment of patients with Covid-19 with a high dose of ulinastatin.

Currently, there are no specific therapeutic agents available for the treatment of coronavirus disease 2019 (Covid-19). The present study aimed to assess the efficacy of high-dose ulinastatin for the treatment of patients with Covid-19. A total of 12 patients hospitalized with confirmed severe acute respiratory syndrome coronavirus 2 infection were treated with a high dose of ulinastatin alongside standard care. Changes in clinical manifestations, laboratory examinations and chest images were retrospectively analyzed. A total of 10 patients with severe Covid-19 and two patients with moderate Covid-19 received ulinastatin treatment. The average age of the patients was 68.0±11.9 years (age range, 48-87 years). In total, nine of the 12 patients (75.0%) had one or more comorbidities. The most common symptoms on admission were fever (8/12, 66.7%), cough (5/12, 41.7%) and dyspnea (5/12, 41.7%). The percentage of lymphocytes was decreased in 41.7% of patients (5/12) and 58.3% of patients (7/12) had elevated hypersensitive C-reactive protein (CRP) levels (mean, 49.70±77.70 mg/l). The white blood cell levels and the percentage of lymphocytes returned to normal in all of the patients, and CRP was significantly decreased and returned to normal in 83.3% of patients (10/12; mean, 6.87±6.63 mg/l) on day 7 after ulinastatin treatment. Clinical symptoms were relieved synchronously. The peripheral oxygen saturation improved and 66.7% of the patients (8/12) did not require further oxygen therapy 7 days after ulinastatin treatment. No patients required intensive care unit admission or mechanical ventilation. All patients revealed different degrees of absorption of pulmonary lesions after treatment. Compared with the standard care group, ulinastatin treatment significantly prevented illness deterioration. In conclusion, these preliminary data revealed that high-dose ulinastatin treatment was safe and exhibited a potential beneficial effect for patients with Covid-19.

The presence of NAFLD in nonobese subjects increased the risk of metabolic abnormalities than obese subjects without NAFLD: a population-based cross-sectional study.

BACKGROUND: With lifestyle modification and over-nutrition, the prevalence of nonalcoholic fatty liver disease (NAFLD) has been increasing annually. Here we aimed to assess the updated prevalence of NAFLD, and to evaluate the association of NAFLD with metabolic abnormalities according to gender, body mass index and age. METHODS: A population-based cross-sectional study was conducted in Shanghai from December 2016 to July 2017. With a three-stage stratified sampling strategy, 3,717 eligible participants were enrolled for the analysis. RESULTS: In total, 1,217 subjects (32.7%) had NAFLD. Among them, 400 (16.3%) of the nonobese and 817 (65.0%) of the obese subjects had NAFLD. The prevalence of NAFLD was increased according to the quartiles of age and waist circumference (WC) in the nonobese subjects. Females with nonobese NAFLD had 1.6-, 2.6-, 2.0-, 2.3- and 3.3-fold higher risks for metabolic syndrome, diabetes mellitus, hyperglycemia, hypertriglycerdemia (high TG) and low high-density lipoprotein cholesterol than obese subjects without NAFLD, respectively. Males had comparable metabolic profiles in both groups, except for a 2.0-fold higher risk of high TG in nonobese NAFLD subjects compared with obese subjects without NAFLD. More impressively, the homeostasis metabolic assessment insulin resistance index was comparable between the two groups. CONCLUSIONS: The increase of age and WC had significant impact on the risk of NAFLD in nonobese subjects. The presence of NAFLD in nonobese subjects increased the risk of metabolic diseases than obese subjects without NAFLD, especially in female.

SHP-1 ameliorates nonalcoholic steatohepatitis by inhibiting proinflammatory cytokine production.

Inflammation is the main contributor for the pathogenesis of nonalcoholic steatohepatitis (NASH). Src homology region 2 domain-containing phosphatase 1 (SHP-1, also known as PTPN6) is regarded as a negative regulator of inflammation, but its role in NASH remains unknown. Here, hepatocyte-specific Ptpn6 knockout mice (Ptpn6HKO ) and adenovirus vector-mediated ectopic expression of SHP-1 (AdSHP1) were used to evaluate the role of SHP-1 in a methionine- and choline-deficient diet-induced NASH model. Compared with the control littermates, Ptpn6HKO mice show exacerbated hepatic steatosis, inflammation, and fibrosis. Additionally, administration of AdSHP1 significantly ameliorates steatohepatitis and inhibits the expression of proinflammatory cytokines, including transforming growth factor-ß, interleukin-6, and tumor necrosis factor-α. Our data indicate that SHP-1 could be a potential therapeutic target for NASH.

Association study between genetic variants in vitamin D metabolism related genes and childhood autism spectrum disorder.

Vitamin D deficiency has been implicated as a risk factor for autism spectrum disorder (ASD). This case-controlled study was to determine the association between single nucleotide polymorphisms (SNPs) in genes encoding vitamin D metabolism related enzymes and childhood ASD in a Chinese Han population. Both autistic children and age-and gender-matched healthy controls were recruited from September 2012-November 2017. The severity of ASD was evaluated by the childhood autism rating scale (CARS). Taqman probe based real-time PCR was applied to examine genotypes. The association between SNPs and the risk of ASD or the disease severity was examined through the logistic regression. This study recruited 249 children with ASD and 353 healthy controls. The G/A genotype (P = 0.0112) or the G allele (P = 0.0117) of CYP24A1 rs17219315, and the G/A genotype of CYP27B1 rs4646536 (P = 0.0341) were significantly associated with an increased risk of ASD. In addition, multivariate analysis found that A allele of both CYP2R1 rs12794714 (P = 0.0159) and CYP27B1 rs4646536 (P = 0.0268) were significantly associated with the severity of ASD. Genetic polymorphisms in vitamin D metabolism related enzymes are associated with the risk of childhood ASD and the severity of the disease.

MicroRNA-134 deactivates hepatic stellate cells by targeting TGF-ß activated kinase 1-binding protein 1.

Aberrant expression of microRNAs is associated with liver fibrogenesis. We previously found that microRNA-134 (miR-134) expression was reduced in fibrosis-based hepatocarcinogenesis induced by diethylinitrosamine. Herein we investigate the role and mechanisms of miR-134 in hepatic fibrosis. Our data show that miR-134 expression is reduced in rat hepatic fibrogenesis induced by carbontetrachloride, bile duct ligation, and dimethylnitrosamine, as well as in activated hepatic stellate cells (HSCs). Moreover, miR-134 inhibited HSC proliferation, and decreased the expression of smooth muscle actin and collagen I in HSCs, whereas the miR-134 inhibitor increased HSC activation. MiR-134 also negatively regulated transforming growth factor-ß-activated kinase 1-binding protein 1 (TAB1) expression in both human and rat HSCs by directly binding to its 3' untranslated region. Importantly, TAB1 expression was significantly elevated during liver fibrogenesis and HSC activation. Knockdown of TAB1 inhibited the proliferation and fibrogenic behavior of HSCs, and significantly reduced the effect of the miR-134 inhibitor on HSC proliferation. Collectively, these data suggest that miR-134 inhibits the activation of HSCs via directly targeting TAB1, and the restoration of miR-134 or targeting TAB1 is of clinical significance in the treatment of liver fibrosis.

Long noncoding RNA C21orf121/bone morphogenetic protein 2/microRNA-140-5p gene network promotes directed differentiation of stem cells from human exfoliated deciduous teeth to neuronal cells.

Previous studies have revealed that long noncoding RNA (lncRNA) and microRNA play a crucial role in autism, which is a childhood neurodevelopmental disorder with complicated genetic origins. Hence, the study concerns whether lncRNA C21orf121/bone morphogenetic proteins 2 (BMP2)/miR-140-5p gene network affects directed differentiation of stem cells from human exfoliated deciduous teeth (SHED) to neuronal cells in rats with autism. Autism models were successfully established. The neuron cells that differentiated from SHED cell were identified. The expression of lncRNA C21orf121, miR-140-5p, BMP2, Nestin, ßIII-tubulin, and microtubule-associated protein 2 (MAP2) and the expression of neuron-specific enolase (NSE) were examined. Besides, the gap junction (GJ) function of SHED, the intracellular free Ca 2+ concentration, and the social behavior and repetitive stereotyped movements of rats in autism were detected. The target relationship between lncRNA C21orf121 and miR-140-5p and that between miR-140-5p and BMP2 were also verified. Firstly, we successfully isolated SHED and identified the differentiated neurons of SHED. Besides, the expression of BMP2, MAP2, Nestin, ßIII-tubulin, NSE positive rate, GJ function, and intracellular free Ca 2+ concentration were increased with the upregulation of C21orf121 and downregulation of miR-140-5p, and accumulated time of repetitive stereotyped movements decreased and the frequency of social behavior increased. The results indicate that lncRNA C21orf121 as a competing endogenous RNA competes with BMP2 binding to miR-140-5p, thereby promoting SHED to differentiate into neuronal cells via upregulating BMP2 expression.

SHP-1 Acts as a Tumor Suppressor in Hepatocarcinogenesis and HCC Progression.

Src homology region 2 (SH2) domain-containing phosphatase 1 (SHP-1, also known as PTPN6) is a nonreceptor protein tyrosine phosphatase that acts as a negative regulator of inflammation. Emerging evidence indicates that SHP-1 plays a role in inhibiting the progression of hepatocellular carcinoma (HCC). However, the role of SHP-1 in hepatocarcinogenesis remains unknown. Here, we find that levels of SHP-1 are significantly downregulated in human HCC tissues compared with those in noncancerous tissues (P < 0.001) and inversely correlate with tumor diameters (r = -0.4130, P = 0.0002) and serum α-fetoprotein levels (P = 0.047). Reduced SHP-1 expression was associated with shorter overall survival of patients with HCC with HBV infection. Overexpression of SHP-1 suppressed proliferation, migration, invasion, and tumorigenicity of HCC cells, whereas knockdown of SHP-1 enhanced the malignant phenotype. Moreover, knockout of Ptpn6 in hepatocytes (Ptpn6HKO ) enhanced hepatocarcinogenesis induced by diethylnitrosamine (DEN) as well as metastasis of primary liver cancer in mice. Furthermore, systemic delivery of SHP-1 by an adenovirus expression vector exerted a therapeutic effect in an orthotopic model of HCC in NOD/SCID mice and DEN-induced primary liver cancers in Ptpn6HKO mice. In addition, SHP-1 inhibited the activation of JAK/STAT, NF-κB, and AKT signaling pathways, but not the MAPK pathway in primary hepatocytes from DEN-treated mice and human HCC cells. Together, our data implicate SHP-1 as a tumor suppressor of hepatocarcinogenesis and HCC progression and propose it as a novel prognostic biomarker and therapeutic target of HCC.Significance: The nonreceptor protein tyrosine phosphatase SHP-1 acts as a tumor suppressor in hepatocellular carcinoma. Cancer Res; 78(16); 4680-91. ©2018 AACR.

Small intestinal hemangioma: Endoscopic or surgical intervention? A case report and review of literature.

BACKGROUND: Hemangioma of the small intestine is a rare vascular malformation. Before the advent of capsule endoscopy (CE) and balloon-assisted enteroscopy (BAE), preoperative diagnosis of this disease was extremely difficult. CASE SUMMARY: In this study, we report a 24-year-old female with a large transmural small bowel cavernous hemangioma, which was diagnosed with CE and BAE preoperatively and removed successfully using minimally invasive surgery. Meanwhile, we perform a literature review of the studies about intestinal hemangiomas published after 2000. Literature review revealed that 91.9% of the lesions were diagnosed preoperatively by CE and/or BAE and 45.9% of them were treated endoscopically, which is a marked improvement compared to before 2000. Therefore, CE and BAE are useful modalities for the preoperative diagnosis of hemangiomas in the small intestine. CONCLUSION: Endoscopic treatment of intestinal hemangioma is generally prudent and might be suitable for multiple, relatively small lesions.

A study of single nucleotide polymorphisms in CD157, AIM2 and JARID2 genes in Han Chinese children with autism spectrum disorder.

PURPOSE: Autism spectrum disorder (ASD) is a group of developmental brain disorders caused by genetic and environmental factors. The objective of this study was to investigate whether single nucleotide polymorphisms (SNPs) in genes related to immune function were associated with ASD in Chinese Han children. MATERIALS AND METHODS: A total of 201 children with ASD and 200 age- and gender-matched healthy controls were recruited from September 2012 to June 2106. A TaqMan probe-based approach was used to genotype SNPs corresponding to rs28532698 and rs4301112 in CD157, rs855867 in AIM2, and rs2237126 in JARID2. Case-control and case-only studies were performed to determine the contribution of SNPs to the predisposition of disease and its severity, respectively. RESULTS: Our results revealed that the genotypes and allele frequencies of these SNPs were not significantly associated with childhood ASD and its severity in this population. CONCLUSIONS: Results of our study suggest that these SNPs are not predictors of childhood ASD in the Chinese Han population. The discrepant results suggest the predictor roles of SNPs have to be determined in different ethnic populations due to genetic heterogeneity of ASD.

Single Nucleotide Polymorphisms in SLC19A1 and SLC25A9 Are Associated with Childhood Autism Spectrum Disorder in the Chinese Han Population.

Genetic variants have been implicated in the development of autism spectrum disorder (ASD). Recent studies suggest that solute carriers (SLCs) may play a role in the etiology of ASD. This purpose of this study was to determine the association between single nucleotide polymorphisms (SNPs) in SLC19A1 and SLC25A12 genes with childhood ASD in a Chinese Han population. A total of 201 autistic children and 200 age- and gender-matched healthy controls were recruited. A TaqMan probe-based real-time PCR approach was used to determine genotypes of SNPs corresponding to rs1023159 and rs1051266 in SLC19A1, and rs2056202 and rs2292813 in SLC25A12. Our results showed that both the T/T genotype of rs1051266 (odds ratio (OR) = 1.85, 95% confidence interval (CI) = 1.06-3.23, P = 0.0301) and the T allele (OR = 1.77, 95% CI = 1.07-2.90, P = 0.0249) of rs2292813 were significantly associated with an increased risk of childhood ASD. In addition, the G-C haplotype of rs1023159-rs1051266 in SCL19A1 (OR = 0.71, 95% CI = 0.51-0.98, P = 0.0389) and C-C haplotype of rs2056202-rs2292813 in SLC25A12 (OR = 0.58, 95% CI = 0.35-0.96, P = 0.0325) were associated with decreased risks of childhood ASD. There was no significant association between genotypes and allele frequencies with the severity of the disease. Our study suggests that these genetic variants of SLC19A1 and SLC25A12 may be associated with risks for childhood ASD.

Silencing of plasminogen activator inhibitor-1 suppresses colorectal cancer progression and liver metastasis.

BACKGROUND: Plasminogen activator inhibitor-1 (PAI-1) is reported to be expressed in many cancer cell types and regarded as one of the most informative biochemical markers for poor prognosis. However, no previous study has evaluated whether PAI-1 could serve as a target in antitumor and antimetastasis therapies of colorectal cancer (CRC). METHODS: The plasma level of PAI-1 in CRC patients was detected and its correlation with the clinicopathologic features was evaluated. PAI-1 protein expression was assessed by Western blot assay and immunohistochemistry. The biologic consequences of PAI-1 silencing in colon cancer cell lines and CRC bearing nude mice were also investigated. RESULTS: Plasma PAI-1 level was higher in CRC patients with liver metastasis and correlated with liver metastasis, tumor size, differentiation, serosa infiltration, Duke's stage, and lymphatic metastasis. PAI-1 protein expression in the CRC tissue of patients with liver metastasis was significantly greater than that in those without liver metastasis. In addition, the abilities of proliferation, invasion, and migration of CRC cells transfected with lentivirus expressing PAI-1 small interfering RNA were reduced significantly. Nude mice inoculated with PAI-1 knockdown cells also had fewer metastatic nodules in the liver and smaller tumor volumes. CONCLUSION: Plasma PAI-1 level was increased in CRC patients with liver metastasis, and PAI-1 silencing may significantly compromise the malignant behaviors of CRC cells in vitro and in vivo. These findings may provide evidence for PAI-1 targeted therapy of CRC.

The transcription factor FOXA2 suppresses gastric tumorigenesis in vitro and in vivo.

BACKGROUND AND AIMS: The transcription factor forkhead box A2 (FOXA2) plays a central role in the development of endoderm-derived organs. It has been reported that FOXA2 acts as a suppressor in many kinds of tumor. However, little is known about the role of FOXA2 in gastric cancer. METHODS: The expression of FOXA2 in gastric cancer tissue samples from 89 patients was assessed by immunohistochemistry, and the clinicopathological characteristics of the samples were analyzed. The human gastric cancer cell line, BGC-823, was used to investigate the effects of FOXA2 in gastric cancer in vitro and in vivo and the potential mechanism involved was explored. RESULTS: FOXA2 expression in human gastric cancer cell lines and human gastric cancer tissues was lower compared with the normal gastric epithelium cell line GES1 and normal adult gastric tissues, respectively. Patients with high FOXA2 expression level had longer 5-year overall survival than those with low FOXA2 expression level. FOXA2 markedly inhibited growth of BGC-823 cells accompanied with the cell cycle arrest and apoptosis. Infection of BGC-823 cells by FOXA2 lentivirus resulted in reduced cell tumorigenesis in vitro and in vivo. Moreover, expression of Mucin 5AC was up-regulated along with increased expression of exogenous FOXA2 in BGC-823 cells; in contrast, dedifferentiation markers, BMI, CD54 and CD24, were down-regulated. CONCLUSIONS: These results suggest that FOXA2 induces the differentiation of gastric cancer and highlight FOXA2 as a novel therapeutic target and prognostic marker for human gastric cancer.

FOXA2 suppresses the metastasis of hepatocellular carcinoma partially through matrix metalloproteinase-9 inhibition.

The forkhead box transcription factor A2 (FOXA2) is a member of the hepatocyte nuclear factor family and plays an important role in liver development and metabolic homeostasis, but its role in the metastasis of hepatocellular carcinoma (HCC) has not been evaluated. In this study, we found that the expression of FOXA2 was decreased in 68.1% (49/72) of human HCC tissues compared with their paired non-cancerous adjacent tissues. Clinicopathological analysis revealed that reduced FOXA2 expression was correlated with aggressive characteristics (venous invasion, poor differentiation, high tumor node metastasis grade). FOXA2 level was even lower in portal vein tumor thrombus compared with primary tumor tissues and correlated with epithelial-mesenchymal transition in HCC cells. Overexpression of FOXA2 inhibited migration and invasion of Focus cells, whereas knockdown of FOXA2 in HepG2 showed the opposite effect. Moreover, upregulation of FOXA2 suppressed HCC metastasis to bone, brain and lung in two distinct mouse models. Finally, we proved that FOXA2 repressed the transcription of matrix metalloproteinase (MMP)-9 and exerted its antimetastasis effect partially through downregulation of MMP-9. In conclusion, our findings indicate that FOXA2 plays a critical role in HCC metastasis and may serve as a novel therapeutic target for HCC.

Induced neural stem cells generated from rat fibroblasts.

The generation of induced tissue-specific stem cells has been hampered by the lack of well-established methods for the maintenance of pure tissue-specific stem cells like the ones we have for embryonic stem (ES) cell cultures. Using a cocktail of cytokines and small molecules, we demonstrate that primitive neural stem (NS) cells derived from mouse ES cells and rat embryos can be maintained. Furthermore, using the same set of cytokines and small molecules, we show that induced NS (iNS) cells can be generated from rat fibroblasts by forced expression of the transcriptional factors Oct4, Sox2 and c-Myc. The generation and long-term maintenance of iNS cells could have wide and momentous implications.

Sophocarpine alleviates non-alcoholic steatohepatitis in rats.

BACKGROUND AND AIM: Non-alcoholic steatohepatitis (NASH) is one entity in the spectrum of non-alcoholic fatty liver disease (NAFLD). The aim of this study was to explore the prevention and therapeutic effect of sophocarpine on experimental rat NASH. METHODS: Sophocarpine with the dosage of 20 mg/kg/day was injected into NASH rats. At the end of 12 weeks, all rats were killed to detect the degree of fatty degeneration, inflammation and fibrosis. RESULTS: Sophocarpine intervention (in the pro-treated and treated groups) resulted in a significant decrease of liver weight, liver index, serum transaminase and serum lipids. Messenger RNA expressions of leptin, interleukin (IL)-6, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-ß1, procollagen-I and α-smooth muscle actin (SMA) and deposition of IL-6, TNF-α and TGF-ß1 in liver decreased, whereas the messenger RNA expression of adiponectin increased significantly compared with that in the model group. Moreover, histological improvement was also observed in the sophocarpine intervention group. In addition, there was no significant difference in any detected indicator between the pro-treated and treated group. CONCLUSIONS: Sophocarpine could decrease the level of serum transaminase, improve lipid metabolism, reduce synthesis of inflammatory cytokines TNF-α, TGF-ß1 and IL-6, activate protective adipocytokine adiponectin, and might be selected as a promising agent for the clinical prevention and therapy of NASH.

Inhibition of extracellular signal-regulated kinase 1 by adenovirus mediated small interfering RNA attenuates hepatic fibrosis in rats.

UNLABELLED: Extracellular signal-regulated kinase 1 (ERK1) is a critical part of the mitogen-activated protein kinase signal transduction pathway, which is involved in hepatic fibrosis. However, the effect of down-regulation of ERK1 on hepatic fibrosis has not been reported. Here, we induced hepatic fibrosis in rats with dimethylnitrosamine administration or bile duct ligation. An adenovirus carrying small interfering RNA targeting ERK1 (AdshERK1) was constructed to determine its effect on hepatic fibrosis, as evaluated by histological and immunohistochemical examination. Our results demonstrated that AdshERK1 significantly reduced the expression of ERK1 and suppressed proliferation and levels of fibrosis-related genes in hepatic stellate cells in vitro. More importantly, selective inhibition of ERK1 remarkably attenuated the deposition of the extracellular matrix in fibrotic liver in both fibrosis models. In addition, both hepatocytes and biliary epithelial cells were proven to exert the ability to generate the myofibroblasts depending on the insults of the liver, which were remarkably reduced by AdshERK1. Furthermore, up-regulation of ERK1 paralleled the increased expression of transforming growth factor beta1 (TGF-beta1), vimentin, snail, platelet-derived growth factor-BB (PDGF-BB), bone morphogenetic protein 4 (BMP4), and small mothers against decapentaplegic-1 (p-Smad1), and was in reverse correlation with E-cadherin in the fibrotic liver. Nevertheless, inhibition of ERK1 resulted in the increased level of E-cadherin in parallel with suppression of TGF-beta1, vimentin, snail, PDGF-BB, BMP4, and p-Smad1. Interestingly, AdshERK1 treatment promoted hepatocellular proliferation. CONCLUSION: Our study provides the first evidence for AdshERK1 suppression of hepatic fibrosis through the reversal of epithelial-mesenchymal transition of both hepatocytes and biliary epithelial cells without interference of hepatocellular proliferation. This suggests that ERK1 is implicated in hepatic fibrogenesis and selective inhibition of ERK1 by small interfering RNA may present a novel option for hepatic fibrosis treatment.

Targeted RNA interference for hepatic fibrosis.

Hepatic fibrosis is a common consequence in patients with chronic liver damage. To date, no agent has been approved for the treatment of hepatic fibrosis. RNA interference (RNAi) is known to be a powerful tool for post-transcriptional gene silencing and has opened new avenues in gene therapy. The problems of lack of cell specificity in vivo and subsequently the occurrence of side effects has hampered the development of hepatic fibrosis treatment. To overcome these shortcomings, several targeted strategies have been developed, such as hydrodynamics-based approaches, local administration, cell-type-selective ligands and cell-type-specific promoters or enhancers, etc. Here, we provide an overview of targeted strategies for the treatment of hepatic fibrosis, and particularly, targeted RNAi for hepatic fibrosis.

Adenovirus-mediated transfer of siRNA against PAI-1 mRNA ameliorates hepatic fibrosis in rats.

BACKGROUND/AIMS: Plasminogen activator inhibitor-1 (PAI-1) is a potential profibrotic molecule. The aim of this study was to evaluate the therapeutic effect of PAI-1 small interfering RNA (siRNA) on experimental hepatic fibrosis and investigate the intrinsic mechanisms. METHODS: Hepatic fibrosis in rats was induced by dimethylnitrosamine (DMN) administration or bile duct ligation (BDL). An adenovirus carrying PAI-1 shRNA (AdshPAI) was generated and administered via tail vein injection. The expression of PAI-1 was confirmed by real-time RT-PCR and immunohistochemistry. The effect of AdshPAI on fibrosis was evaluated by histological and immunohistochemical examination. RESULTS: We found that PAI-1 was downregulated after AdshPAI administration. Liver fibrosis was significantly improved after AdshPAI administration in both DMN and BDL models. AdshPAI treatment facilitated matrix degradation by correcting the levels of matrix metalloproteinases (MMPs) and its inhibitors (TIMPs) through upregulation of MMP9, MMP13 and downregulation of TIMP-1. Moreover, AdshPAI treatment stimulated hepatocellular proliferation and inhibited cellular apoptosis. CONCLUSIONS: This study suggests that AdshPAI treatment has a protective effect on hepatocytes and ameliorates liver fibrogenesis. Inhibiting the upregulation of PAI-1 during liver fibrosis may be an antifibrotic pathway worth exploiting.

Inhibition of plasminogen activator inhibitor-1 expression by siRNA in rat hepatic stellate cells.

BACKGROUND AND AIM: The plasminogen activator/plasmin system is known to regulate the extracellular matrix turnover. The aim of this study was to detect the role of plasminogen activator inhibitor-1 (PAI-1) during liver fibrogenesis and investigate the functional effects of PAI-1 gene silencing in rat hepatic stellate cells (HSCs) using small interfering RNA (siRNA). METHODS: Hepatic fibrosis in rats was induced through serial subcutaneously injections of CCl(4) and the expression of PAI-1 was detected by immunohistochemistry and reverse transcription-polymerase chain reaction (PCR). PAI-1 siRNA molecules were constructed and transiently transfected into HSC-T6 using the cell suspension transfection method. The pSUPER RNA interfering system was used to establish the HSC stable cell line pSUPER-shPAI. Expression of alpha-smooth muscle actin, transforming growth factor-beta, tissue inhibitor of metalloproteinases-1, and collagen types I and III were evaluated by real-time PCR. Cell proliferation and the cell cycle were determined by the methyl thiazolyl tetrazolium (MTT) method and flow cytometry. Collagen content in HSCs supernatant was evaluated by enzyme-linked immunosorbent assay. RESULTS: The results showed that PAI-1 was upregulated during liver fibrosis, and its expression was closely correlated with the deposition of collagens. SiRNA molecules were successfully transfected into HSCs and induced inhibition of PAI-1 expression time dependently. Moreover, PAI-1 siRNA treatment downregulated alpha-smooth muscle actin, transforming growth factor-beta, tissue inhibitor of metalloproteinases-1 expression, and inhibited collagen types I and III synthesis both at the mRNA and protein level in transiently and stably transfected HSCs. CONCLUSIONS: This study suggests a significant functional role for PAI-1 in the development of liver fibrosis and that downregulating PAI-1 expression might present as a potential strategy to treat liver fibrosis.