Clinical and genetic characterization of pediatric patients with progressive familial intrahepatic cholestasis type 3 (PFIC3): identification of 14 novel ABCB4 variants and review of the literatures.

BACKGROUND: Progressive familial intrahepatic cholestasis type 3 (PFIC3) is an autosomal recessive disease caused by pathogenic variants of the gene ABCB4. This study aimed to investigate the ABCB4 genotypic and the clinical phenotypic features of PFIC3 patients. METHODS: The clinical and molecular genetic data of 13 new pediatric patients with PFIC3 as well as 82 reported ones in the PubMed and CNKI databases were collected and analyzed. RESULTS: The 13 new PFIC3 patients included six females and seven males, and the main presentations were hepatomegaly, splenomegaly, jaundice, and pruritus, as well as increased levels of gamma-glutamyl transpeptidase (GGT). Fourteen new ABCB4 variants were detected, including eight diagnosed to be likely-pathogenic and six, pathogenic. Among all the 95 PFIC3 cases, hepatomegaly was observed in 85.3% (81/95), pruritus in 67.4% (64/95), splenomegaly in 52.6% (50/95), jaundice in 48.4% (46/95), portal hypertension in 34.7% (33/95) and GGT elevation in 100% (88/88) of the patients. Positive responses at varied degrees to oral ursodeoxycholic acid (UDCA) treatment were observed in 66.1% (39/59) of the patients, among whom 38.5% (15/39) fully recovered in terms of the laboratory changes. Although the condition remained stable in 53 patients (58.9%, 53/90), the clinical outcomes were not promising in the rest 37 cases (41.1%, 37/90), including 7 died, 27 having undergone while another 3 waiting for liver transplantation. A total of 96 ABCB4 variants were detected in the 95 patients. PFIC3 patients with biallelic null variants exhibited earlier onset ages [10.5 (2, 18) vs. 19 (8, 60) months, p = 0.007], lower UDCA response rate [18.2% (2/11) vs. 77.1% (37/48), p = 0.001], and more unpromising clinical outcomes [80% (12/15) vs. 33.3% (25/75), p = 0.001], compared with those with non-biallelic null variants. CONCLUSIONS: PFIC3 presented with hepatomegaly, pruritus, splenomegaly and jaundice with increased serum GGT level as a biochemistry hallmark. Although varying degrees of improvement in response to UDCA therapy were observed, 41.1% of PFIC3 patients exhibited unfavorable prognosis. ABCB4 genotypes of biallelic null variants were associated with severer PFIC3 phenotypes. Moreover, the 14 novel variants in this study expanded the ABCB4 mutation spectrum, and provided novel molecular biomarkers for diagnosis of PFIC3 patients.

MiR-125b enhances autophagic flux to improve septic cardiomyopathy via targeting STAT3/HMGB1.

We explore the role of miR-125b in septic cardiomyopathy, focusing on miR-125b/STAT3/HMGB1 axis. CLP mouse model and LPS-stimulated primary rat cardiomyocytes (CMs) and H9C2 cell were used as in vivo and in vitro models of septic cardiomyopathy, respectively. qRT-PCR and western blot were performed to measure expression levels of miR-125b, STAT3, HMGB1, and autophagy-related proteins. MTT assay was employed to examine LPS toxicity. Dual luciferase activity assay and CHIP were performed to validate interactions of miR-125b/STAT3 and STAT3/HMGB1 promoter. Immunostaining was used to assess the level of autophagic flux. ROS level was measured by fluorescence assay. Heart functions were examined via intracoronary Doppler ultrasound. miR-125b was diminished while STAT3 and HMGB1 were elevated in the heart tissue following CLP surgery and in LPS-treated H9C2 cells. LPS treatment up-regulated ROS generation and suppressed autophagic flux. Overexpression of miR-125b mimics or knockdown of STAT3 or HMGB1 alleviated LPS-induced hindrance of autophagic flux and ROS production. miR-125b directly targeted STAT3 mRNA and STAT3 bound with HMGB1 promoter. Overexpression of miR-125b mitigated myocardial dysfunction induced by CLP in vivo. Hyperactivation of STAT3/HMGB1 caused by reduced miR-125b contributes to ROS generation and the hindrance of autophagic flux during septic cardiomyopathy, leading to myocardial dysfunction.

CircTLK1 modulates sepsis-induced cardiomyocyte apoptosis via enhancing PARP1/HMGB1 axis-mediated mitochondrial DNA damage by sponging miR-17-5p.

INTRODUCTION: Septic cardiomyopathy is a common complication of sepsis with high morbidity and mortality, but lacks specific therapy. This study aimed to reveal the role of circTLK1 and its potential mechanisms in septic cardiomyopathy. MATERIALS AND METHODS: The in vitro and in vivo models of septic cardiomyopathy were established. Cell viability and apoptosis were detected by CCK8, TUNEL and flow cytometry, respectively. LDH, CK, SOD, MDA, ATP, 8-OHdG, NAD+/NADH ratio, ROS level, mitochondrial membrane potential and cytochrome C distribution were evaluated using commercial kits. qRT-PCR and western blotting were performed to detect RNA and protein levels. Mitochondrial DNA (mtDNA) copy number and transcription were assessed by quantitative PCR. Dual-luciferase assay, RNA immunoprecipitation and co-immunoprecipitation were performed to verify the interaction between circTLK1/PARP1 and miR-17-5p. RESULTS: CircTLK1, PARP1 and HMGB1 were up-regulated in the in vitro and in vivo models of septic cardiomyopathy. CircTLK1 inhibition restrained LPS-induced up-regulation of PARP1 and HMGB1. Moreover, circTLK1 knockdown repressed sepsis-induced mtDNA oxidative damage, mitochondrial dysfunction and consequent cardiomyocyte apoptosis by inhibiting PARP1/HMGB1 axis in vitro and in vivo. In addition, circTLK1 enhanced PARP1 expression via sponging miR-17-5p. Inhibition of miR-17-5p abolished the protective effects of circTLK1 silencing on oxidative mtDNA damage and cardiomyocyte apoptosis. CONCLUSION: CircTLK1 sponged miR-17-5p to aggravate mtDNA oxidative damage, mitochondrial dysfunction and cardiomyocyte apoptosis via activating PARP1/HMGB1 axis during sepsis, indicating that circTLK1 may be a putative therapeutic target for septic cardiomyopathy.

Blockchain Personal Health Records: Systematic Review.

BACKGROUND: Blockchain technology has the potential to enable more secure, transparent, and equitable data management. In the health care domain, it has been applied most frequently to electronic health records. In addition to securely managing data, blockchain has significant advantages in distributing data access, control, and ownership to end users. Due to this attribute, among others, the use of blockchain to power personal health records (PHRs) is especially appealing. OBJECTIVE: This review aims to examine the current landscape, design choices, limitations, and future directions of blockchain-based PHRs. METHODS: Adopting the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-analyses) guidelines, a cross-disciplinary systematic review was performed in July 2020 on all eligible articles, including gray literature, from the following 8 databases: ACM, IEEE Xplore, MEDLINE, ScienceDirect, Scopus, SpringerLink, Web of Science, and Google Scholar. Three reviewers independently performed a full-text review and data abstraction using a standardized data collection form. RESULTS: A total of 58 articles met the inclusion criteria. In the review, we found that the blockchain PHR space has matured over the past 5 years, from purely conceptual ideas initially to an increasing trend of publications describing prototypes and even implementations. Although the eventual application of blockchain in PHRs is intended for the health care industry, the majority of the articles were found in engineering or computer science publications. Among the blockchain PHRs described, permissioned blockchains and off-chain storage were the most common design choices. Although 18 articles described a tethered blockchain PHR, all of them were at the conceptual stage. CONCLUSIONS: This review revealed that although research interest in blockchain PHRs is increasing and that the space is maturing, this technology is still largely in the conceptual stage. Being the first systematic review on blockchain PHRs, this review should serve as a basis for future reviews to track the development of the space.

ß-catenin-coordinated lncRNA MALAT1 up-regulation of ZEB-1 could enhance the telomerase activity in HGF-mediated differentiation of bone marrow mesenchymal stem cells into hepatocytes.

OBJECTIVE: To investigate role of ß-catenin and lncRNA MALAT1/miR-217 axis to converge into the regulation of ZEB-1 in hepatocyte growth factor (HGF)-induced hepatocytes differentiated from bone marrow mesenchymal stem cells (BM-MSCs). METHODS: BM-MSCs were isolated and HGF was used to induce the differentiation of BM-MSCs into hepatocytes. HSC-T6 cells, BRL-3 A cells and differentiated BM-MSCs were treated by lipopolysaccharide(LPS). shRNAs were used to silence ß-catenin and recombinant plasmids were used to over-express ZEB1. Measurement of cell viability was conducted using MTT assay and Hoechst 33342 staining. RNA immunoprecipitation (RIP) assay was used to determine binding of miR-217-3p and MALAT1. RESULTS: BM-MSCs successfully differentiated into hepatocytes by HGF treatment. Expression of ß-catenin, ZEB-1 and TERT was up-regulated to a higher level in hepatocytes differentiated from BM-MSCs than HSC-T6 cells and BRL-3 A cells after LPS stimulation. When ß-catenin was knocked down in all cell lines, expression of ß-catenin, ZEB-1 and TERT was significantly decreased as well as telomerase activity. While when ZEB1 was over-expressed, expression of TERT and telomerase activity was all significantly up-regulated. In hepatocytes differentiated from BM-MSCs, miR-217 was down-regulated and lncRNA MALAT1 was up-regulated. RIP analysis showed MALAT1 was physically associated with miR-217 and might function in the regulation of ZEB-1, further enhancing the expression of TERT so as to augment telomerase activity. CONCLUSION: We successfully used HGF to mediate differentiation of BM-MSCs into hepatocytes, and found that ß-catenin-coordinated MALAT1/miR-217 axis could up-regulate expression of ZEB-1 and further enhanced the telomerase activity through regulation of TERT in BM-MSCs differentiating into hepatocytes.

[Changes in Th17 and CD4+CD25+ Treg cells and their significance among children with hand, foot and mouth disease].

OBJECTIVE: To investigate the changes in peripheral blood Th17 and CD4(+)CD25(+) regulatory T (Treg) cells and their significance among children with hand, foot and mouth disease (HFMD). METHODS: Eighty-nine children with HFMD, including 55 cases of common HFMD and 34 cases of severe HFMD, were included in the study; and 30 healthy children were selected as the control group. The percentages of Th17 and CD4(+)CD25(+) Treg cells in CD4(+) T cells in peripheral blood were determined by flow cytometry. The expression levels of interleukin (IL)-10, transforming growth factor-ß (TGF-ß), and IL-17 were measured by enzyme-linked immunosorbent assay. RESULTS: Compared with the control group, the cases of common HFMD and severe HFMD had significantly increased levels of Th17 cells and IL-17 (P<0.05) but significantly decreased levels of CD4(+)CD25(+) Treg cells, IL-10, and TGF-ß (P<0.05). The severity of the HFMD was positively correlated with the levels of Th17 cells and IL-17 in peripheral blood but negatively correlated with the levels of CD4(+)CD25(+) Treg cells, IL-10, and TGF-ß. CONCLUSIONS: Children with HFMD have increased response of Th17 cells but decreased response of CD4(+)CD25(+) Treg cells in peripheral blood. Th17/CD4(+)CD25(+) Treg cell imbalance may play an important role in the pathogenesis of HFMD.

[Pilot observation on the effect of integrated control measures against clonorchiasis in Pengjiang District of Jiangmen City].

From June to December in 2008, five villages were randomly chosen from Pengjiang District of Jiangmen city and about five hundred residents from each village were examined for clonorchiasis by Kato-Katz method (three slides per specimen). Fifty residents from each village were re-examined one month after treatment. One year later 50 treated residents were chosen from Dalin village and Sanya village for fecal examination. Questionnairing was conducted to determine the knowledge rate on clonorchiasis prevention among residents. The percentage and usage of sanitary toilets were investigated. The average infection rate of clonorchiasis from five villages was 21.5%(537/2501). 86.6%(465/537) of clonorchiasis received treatment voluntarily. One month after treatment the infection rate in four villages declined significantly. The positive rate showed no significant difference between one month and one year after treatment in Dalin and Sanya villages (P>0.05) . Questionairing indicated that 41.2%(170/413) of the clonorchiasis cases ate raw fish frequently, which was significantly higher than those non-infected people [4.2%, 8/192] (P<0.05). After health education, the knowledge awareness rate raised from 23.1% (135/584) to 84.5% (349/413) (P<0.05). The dissemination and usage of sanitary toilets were 93.2% (38 068/40 848) and 100%, respectively.

[Role of Treg/Th17 balance in the pathogenesis of cytomegalovirus infection].

AIM: To investigate the role of Treg/Th17 balance in the immune-regulated mechanism of cytomegalovirus (CMV) infection. METHODS: According to the diagnostic criteria of the CMV infection, the patients with CMV infection were divided into the activate infection group and the latent infection group, and the normal children were recruited as control group. The percents of Treg and Th17 in peripheral blood were detected by flow cytometry to calculate Treg/Th17 ratio. ELISA and RT-PCR were performed to determine the levels of the related factors of Treg (IL-10, Foxp3) and Th17 (TGF-ß, IL-17, IL-6, IL-23, ROR-γt), respectively. RESULTS: Compared with the control group, the proportion of Treg in peripheral blood was reduced, Th17 increased, so Treg/Th17 ratio went down after CMV infection (P<0.05). By comparison between two groups of CMV infection, Treg/Th17 ratio and Treg related factors of the activate infection group decreased significantly (P<0.05), but Th17 related factors were up-regulated significantly (P<0.05). CONCLUSION: Treg/Th17 balance takes part in the immune-regulated mechanism of CMV infection, and may be related to virus latent-active state.

[Effects of diallyl disulfide on apoptosis of human leukemia K562 cells and expression of Fas, FasL and caspase-8].

OBJECTIVE: To study the effects of diallyl disulfide (DADS) on apoptosis of human leukemia K562 cells and possible mechanisms. METHODS: The morphologic changes of leukemia K562 cells after DADS treatment were observed by Hoechst 33258 staining. Cell apoptosis rates after different concentrations and different durations of DADS treatment were determined by flow cytometry. Fas, FasL and caspase-8 mRNA expression was estimated by reverse transcription-polymerase chain reaction (RT-PCR) 48 hrs after DADS treatment. RESULTS: The characteristics of apoptosis in K562 cells induced by DADS were observed. After 24 hrs of DADS treatment, the apoptosis rate of K562 cells increased from (11.60 ± 0.83)% at the concentration of 10 mg/L to (37.94 ± 0.87)% at the concentration of 40 mg/L. The apoptosis rate of K562 cells increased after 40 mg/L DADS with the increasing time from (37.94 ± 0.87)% (24 hrs) to (47.02 ± 0.66)% (72 hrs). Expression of Fas and caspase-8 mRNA increased, while FasL mRNA expression decreased significantly 48 hrs after DADS treatment compared with the control group (P<0.05). CONCLUSIONS: DADS can induce apoptosis of human leukemia K562 cells in a time- and concentration-dependent manner, possibly through increasing Fas and caspase-8 expression and decreasing FasL expression.

[Effects of CXCR4 silence induced by RNA interference on cell cycle distribution and apoptosis of Jurkat cells].

This study was aimed to investigate the effect of down-regulating the CXCR4 expression on cell cycle and cell apoptosis of human T-ALL Jurkat cells. The CXCR4 specific siRNA plasmid vector was constructed and then transfected into the cultured Jurkat cell line by DMRIE-C. The expression of CXCR4 mRNA was detected by RT-PCR, the cell distribution in cell cycle and cell apoptosis were determined by flow cytometry. The experiments were divided into 3 groups: group A (blank control), group B (non-silencing dsRNA as negative control) and group C (CXCR4 siRNA). The results showed that the expression level of CXCR4 mRNA in Jurkat cells transfected with CXCR4 siRNA (group C) decreased and cell proportion in G(0)/G(1) phase increased as compared with group A (56.9% +/- 1.4% vs 68.3% +/- 2.4% and 35.8% +/- 1.9% vs 18.1% +/- 1.2% respectively) (p < 0.01), cell proportion in G(2)/M and S phase decreased as compared with group A (19.8% +/- 1.7%, 44.4% +/- 2.1% vs 27.2% +/- 1.5%, 54.7% +/- 2.8% respectively) (p < 0.01). The apoptosis rate of Jurkat cells in group C increased as compared with group A (20.9% +/- 2.0% vs 3.13% +/- 0.9% respectively) (p < 0.01), and the comparison between group A and B showed no statistical difference. It is concluded that the CXCR4 specific siRNA can effectively down-regulate the CXCR4 mRNA expression, which induces the cell apoptosis and cell cycle arrest, thereby inhibits the Jurkat cell proliferation.

[Stem cell factor secretion by bone mesenchymal stem cells stimulated with astragaloside IV].

OBJECTIVE: To study the effect of astragaloside IV on the expression of cytokines in bone mesenchymal stem cells (MSCs) in rats. METHODS: MSCs were isolated from Wistar rats by the method of adhesive cultiration and clone, and then their biological activities were assessed using indirect immunofluorescence. Proliferation of MSCs stimulated with astragaloside IV was ascertained by the MTT method. Expression of cytokines was ascertained using RT-PCR in MSCs with astragaloside IV stimulation or not. RESULTS: MSCs were effectively isolated and purified in vitro, and had expression of many cytokines except IL-3, such as stem cell factor (SCF), thrombopoietin (TPO), granulocyte macrophage colony stimulating factor (GM-CSF) and transforming growth factor (TGF-beta1). Astragaloside IV stimulation promoted MSCs proliferation, and 200 mg/mL astragaloside IV treatment produced a peak effect 72 hrs after culture. The SCF expression in MSCs stimulated with astragaloside IV increased significantly compared with that in MSCs without astragaloside IV stimulation. CONCLUSIONS: Astragaloside IV may promote MSCs proliferation and increase SCF secretion in vitro.

[The role of medical antireflux in asthmatics with gastroesophageal reflux:a systematic review].

OBJECTIVE: to evaluate the effect of antireflux treatment in asthma patients with gastroesophageal reflux disease (GERD). METHODS: prospective randomized trials of antireflux treatment in asthma patients with gastroesophageal reflux were all included. Antireflux treatment should be double-blinded. Sample size and matching ways were not restricted. The study subjects were defined as asthmatic patients with gastroesophageal reflux aged 13 or older. Computer searching included The Cochrane Central Register of Controlled Trials, PubMed, Embase, OVID database, Chinese Biological Medicine database (CBMdisc), China National Knowledge Infrastructure (CNKI), and Wanfang Data. Manual searching included Chinese Journal of Tuberculosis and Respiratory Diseases, Chinese Journal of Digestion, Chinese Journal of Internal Medicine, CHEST and the references of trails included. The search ended on November of 2009. Trials with subjects using antireflux drugs in 3 days before entry, trials repeatly or mutiply published, and trials with methodology quality under grade B were excluded. With the method of Cochrane systematic review, 2 reviewers assessed the quality of the studies, extracted data independently and cross checked the data. Data were combined and analyzed with RevMan 4.3.2 to evaluate the effect of antireflux treatment on asthmatic patients with gastroesophageal reflux disease. RESULTS: fourteen randomized controlled studies met inclusion criteria, including 1555 cases. Meta-analyses demonstrated that antireflux treatment improved the pulmonary function in asthmatics with gastroesophageal reflux. Compared to the placebo group, FEV(1) improved [WMD 0.11 L; 95%CI (0.02 - 0.20); Z = 2.49, P = 0.010]; and PEF also improved, including daytime PEF [WMD 42.33 L/min; 95%CI (3.39 - 81.28), Z = 2.13, P = 0.030], morning PEF [WMD 16.16 L/min; 95%CI (5.91 - 26.41); Z = 3.09, P = 0.002], and nighttime PEF [WMD 18.35 L/min; 95%CI (6.77 - 29.92); Z = 3.11, P = 0.002]. However, no significant difference was found in the decrease of provocative concentration of acetylcholine causing a 20% decrease in FEV(1) (PC20-FEV(1)) [WMD -0.07 mg/L; 95%CI (-0.33 -0.19); Z = 0.55, P = 0.590]. Eight out of 14 studies showed improvement in asthma symptoms after antireflux treatment, but neither daytime symptoms nor nighttime symptoms showed significant improvement in this Meta-analysis. CONCLUSION: antireflux treatment in asthmatics with gastroesophageal reflux could improve pulmonary function, but showed no significant effect on airway hyper-responsiveness or asthma symptoms.