Metabolic Disorders Combined with Noninvasive Tests to Screen Advanced Fibrosis in Nonalcoholic Fatty Liver Disease.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is associated with metabolic disorders. This study aimed to explore the role of metabolic disorders in screening advanced fibrosis in NAFLD patients. METHODS: A total of 246 histologically-proven NAFLD patients were enrolled across 14 centers. We compared the severity of fibrosis in patients with different components of metabolic disorders. Based on standard noninvasive tests and metabolic disorders, we developed new algorithms to identify advanced fibrosis. RESULTS: Metabolic syndrome (MetS) was frequent in NAFLD patients (133/246, 54%). Patients with MetS had a higher proportion of significant fibrosis (p=0.014) and higher LSM values (9.2 kPa, vs. 7.4 kPa, p=0.002) than those without MetS. Patients with more metabolic disorders had higher fibrosis stages (p=0.017). Reduced high-density lipoprotein cholesterol (odds ratio [OR]: 2.241, 95% confidence interval [CI]: 1.004-5.002, p=0.049) and raised fasting glucose (OR: 4.500, 95% CI: 2.083-9.725, p<0.001) were significantly associated with advanced fibrosis. Using these two metabolic disorders as a screening tool, a sensitivity, specificity and accuracy of 92%, 81% and 83% was achieved, respectively. With the new algorithms combining metabolic disorders with noninvasive measurements, the number of patients requiring liver biopsy was reduced, especially in combination with the Fibrosis-4 score and metabolic disorders (36% to 17%, p<0.001). In addition, this stepwise algorithm could achieve a high accuracy (85%) and high negative predictive value (93%). CONCLUSIONS: Metabolic disorders should be taken into consideration in the diagnosis of advanced fibrosis. With further validation and investigation, new algorithms could be recommended in primary care units to spare patients from unnecessary referral and liver biopsies.

The pathologic relevance of metabolic criteria in patients with biopsy-proven nonalcoholic fatty liver disease and metabolic dysfunction associated fatty liver disease: A multicenter cross-sectional study in China.

BACKGROUND: This study aimed to assess the association between metabolic syndrome (MetS) and severity of nonalcoholic fatty liver disease (NAFLD), and to discuss the pathological relevance of the diagnostic criteria in metabolic (dysfunction) associated fatty liver disease (MAFLD). METHODS: This was a multicenter, cross-sectional study. Patients with NAFLD confirmed by liver biopsy were enrolled between July 2016 and December 2018 from 14 centers across the mainland of China. Anthropometric and metabolic parameters were collected to assess the pathological relevance. RESULTS: Of 246 enrolled patients with NAFLD, 150 (61.0%) had the comorbidity of MetS. With the increase of metabolic components, the proportions of nonalcoholic steatohepatitis (NASH) and significant fibrosis were notably increased. The comorbid three metabolic components significantly increased the proportion of NASH, and further increase of metabolic components did not increase the proportion of NASH. However, the increase of metabolic components was parallel to the increase of the proportion of liver fibrosis. Among the 246 patients, 239 (97.2%) met the diagnostic criteria of MAFLD. Although non-MAFLD patients had less NASH, they present with similar proportion of significant fibrosis and cirrhosis. In the diagnostic criteria of MAFLD, BMI ≥ 23 kg/m2 was related to NASH (Mantel-Haenszel Common Estimate OR: 2.975; 95% CI: 1.037-8.538; P = 0.043), and T2DM was related to significant fibrosis (Mantel-Haenszel Common Estimate OR: 2.531; 95% CI: 1.388-4.613; P = 0.002). The homeostasis model assessment of insulin resistance (HOMA-IR) ≥ 2.5 was the most significant factor for NASH (OR: 4.100; 95% CI: 1.772-9.487; P = 0.001) and significant factor for liver fibrosis (OR: 2.947; 95% CI: 1.398-6.210; P = 0.004) after the adjustments of the BMI and diabetes. CONCLUSIONS: Metabolic dysregulations are important risk factors in NAFLD progression. The insulin resistance status may play a predominant role in the progression in MAFLD patients.

Novel compound heterozygous of PARKIN causes early-onset Parkinson's disease.

Genetics has an essential role in the development of early-onset Parkinson's disease (EOPD). Consequently, genetic screening is of great significance for the diagnosis and treatment of EOPD. In this study, we reported two EOPD with compound heterozygous in PARKIN detected by whole-exome sequencing (WES) and ligation-dependent probe amplification (MLPA). Two unrelated EOPD patients and their parents were enrolled in this study. Genetic analysis was performed through WES and verified by direct Sanger sequencing. In addition, MLPA was used to detect exon dosage. Detailed clinical manifestations and several scale assessments were collected for genotype and phenotype analysis. Compound heterozygous mutations in PARKIN were identified in both patients. c.735-1G > A and Ex2del were detected in Case A, while G284R (c.850 G > C) and Ex2del were found in Case B. These variants were confirmed to originate from their normal parents. The c.735-1G > A is a novel PARKIN variant, which was predicted to result from disappearing of the acceptor splice site by NetGene2. The G284R is a previously reported pathological mutation and the Ex2del is a hot variant of PARKIN found in the Asian population. The phenotypes of both patients are quite different, the main manifestation of case A is rigidity onset, while the case B starts with tremor and foot dystonia. In the present study, we reported a novel compound heterozygous form of PARKIN consisting of splice variant c. 735-1G > A and Ex2del. Moreover, we also found that tiny differences in genotypes of PARKIN may lead to obvious clinical phenotypic differences.

Quantifying and monitoring fibrosis in non-alcoholic fatty liver disease using dual-photon microscopy.

OBJECTIVE: Fibrosis stage is strongly associated with liver-related outcomes and is a key surrogate endpoint in drug trials for non-alcoholic steatohepatitis. Dual-photon microscopy allows automated quantification of fibrosis-related parameters (q-FPs) and may facilitate large-scale histological studies. We aim to validate the performance of q-FPs in a large histological cohort. DESIGN: 344 patients with non-alcoholic fatty liver disease (NAFLD) underwent 428 liver biopsies (240 had paired transient elastography examination). Fibrosis stage was scored using the NASH Clinical Research Network system, and q-FPs were measured by dual-photon microscopy using unstained slides. Patients were randomly assigned to the training and validation cohorts to test the performance of individual q-FPs and derive optimal cut-offs. RESULTS: Over 25 q-FPs had area under the receiver-operating characteristics curves >0.90 for different fibrosis stages. Among them, the perimeter of collagen fibres and number of long collagen fibres had the highest accuracy. At the best cut-offs, the two q-FPs had 88.3%-96.2% sensitivity and 78.1%-91.1% specificity for different fibrosis stages in the validation cohort. q-FPs and histological scoring had nearly identical correlations with liver stiffness measurement, suggesting that the accuracy of q-FPs approached that of histological assessment. Among patients with paired liver biopsies, changes in the same q-FPs were associated with changes in fibrosis stage. At a median follow-up of 5.6 years, baseline q-FPs predicted liver-related events. CONCLUSION: q-FP is highly accurate in the assessment of fibrosis in NAFLD patients. This automated platform can be used in future studies as objective and reliable evaluation of histological fibrosis.

Down-regulated expression of microRNA-338-5p contributes to neuropathology in Alzheimer's disease.

Alzheimer's disease (AD) is a leading cause of dementia. However, the mechanisms responsible for development of AD, especially for the sporadic variant, are still not clear. In our previous study, we discovered that a small noncoding RNA (miR-188-3p) targeting ß-site amyloid precursor protein cleaving enzyme (BACE)-1, a key enzyme responsible for Aß formation, plays an important role in the development of neuropathology in AD. In the present study, we identified that miR-338-5p, a new miRNA that also targets BACE1, contributes to AD neuropathology. We observed that expression of miR-338-5p was significantly down-regulated in the hippocampus of patients with AD and 5XFAD transgenic (TG) mice, an animal model of AD. Overexpression of miR-338-5p in the hippocampus of TG mice reduced BACE1 expression, Aß formation, and neuroinflammation. Overexpression of miR-338-5p functionally prevented impairments in long-term synaptic plasticity, learning ability, and memory retention in TG mice. In addition, we provide evidence that down-regulated expression of miR-338-5p in AD is regulated through the NF-κB signaling pathway. Our results suggest that down-regulated expression of miR-338-5p plays an important role in the development of AD.-Qian, Q., Zhang, J., He, F.-P., Bao, W.-X., Zheng, T.-T., Zhou, D.-M., Pan, H.-Y., Zhang, H., Zhang, X.-Q., He, X., Sun, B.-G., Luo, B.-Y., Chen, C., Peng, G.-P. Down-regulated expression of microRNA-338-5p contributes to neuropathology in Alzheimer's disease.

Association of GCKR rs780094 polymorphism with circulating lipid levels in type 2 diabetes and hyperuricemia in Uygur Chinese.

To investigate the relationship between a GCKR rs780094 polymorphism and lipid profiles in the Xinjiang Uygur population in China. 980 type 2 diabetes mellitus (T2DM) patients, 1017 hyperuricemia (HUA) and 1185 healthy controls were included in this study. After genotyping of rs780094 by Sequenom Mass ARRAY system, chi-square test and logistic regression analysis were used for association analysis as well as a genotype-phenotype analysis. We found that the serum concentration of TC (P<0.001) was significantly higher and HDL-C (P<0.001) was lower in T2DM than in control participants. Subjects with HUA had a significantly higher TG (P=0.003) and lower HDL-C (P<0.001) than control participants. Additionally, under the recessive model, rs780094 was shown to be associated with the risk of HUA (P=0.015, OR=1.311), particularly in males (P=0.047, OR=1.330). Subsequent interaction analysis between rs780094 and lipid parameters showed that the TG level was positively correlated with HUA in the rs780094- AA+AG carriers (P=0.005). The TC concentrations showed to be associated with T2DM in the rs780094- AA+AG carriers (P<0.001). The association between lipid parameters and gender showed that significantly higher TG levels (P<0.001) and lower HDL-C levels (P<0.001) were observed in female HUA. Higher LDL-C levels were found in male HUA (P=0.015). Moreover, statistically higher TC levels and lower HDL-C levels were found both in male and female T2DM cases (TC: male: P<0.001, female: P=0.014. HDL-C: male: P<0.001, female: P<0.001.).To conclude, our results demonstrated that different genotypes of rs780094 had different effects on blood lipids in HUA and T2DM patients in a Uygur population. Gender was also one of the factors influencing blood lipid levels.

Correlations between diffusion tensor imaging and levels of consciousness in patients with traumatic brain injury: a systematic review and meta-analysis.

Traumatic brain injury (TBI) often leads to impaired consciousness. Recent diffusion tensor imaging studies associated consciousness with imaging metrics including fractional anisotropy (FA) and apparent diffusion coefficient (ADC). We evaluated their correlations and determined the best index in candidate regions. Six databases were searched, including PubMed and Embase, and 16 studies with 701 participants were included. Data from region-of-interest and whole-brain analysis methods were meta-analysed separately. The FA-consciousness correlation was marginal in the whole-brain white matter (r = 0.63, 95% CI [0.47, 0.79], p = 0.000) and the corpus callosum (CC) (r = 0.60, 95% CI [0.48, 0.71], p = 0.000), and moderate in the internal capsule (r = 0.48, 95% CI [0.24, 0.72], p = 0.000). Correlations with ADC trended negative and lacked significance. Further subgroup analysis revealed that consciousness levels correlated strongly with FA in the CC body (r = 0.66, 95% CI [0.43, 0.89]), moderately in the splenium (r = 0.58, 95% CI [0.38, 0.78]), but insignificantly in the genu. In conclusion, FA correlates better with consciousness levels than ADC in TBI. The degree of correlation varies among brain regions. The CC (especially its splenium and body) is a reliable candidate region to quantitatively reflect consciousness levels.

Protective action of B1R antagonist against cerebral ischemia-reperfusion injury through suppressing miR-200c expression of Microglia-derived microvesicles.

BACKGROUND AND OBJECTIVE: Cerebral ischemia-reperfusion (I/R) injury is a common side-effect for cerebral ischemic disease and its therapeutic regimen is limited. Kinin is pro-inflammatory peptide that is released and acts at the site of injury and inflammation such as brain and it works through bradykinin 1 receptor (B1R). The present study was to examine the effect of B1R antagonist on cerebral I/R injury and the potential mechanism. METHODS: Cerebral I/R injury was induced in mice by transient middle cerebral artery occlusion (MCAO). Neurological function was assessed by Bederson score. Infarct volumes were measured using planimetry. In vitro cell model was made by oxygen-glucose deprivation-Hypoxia/Reoxygenation (OGD-H/R) treatment to N9 microglia cell; and the cultured medium was collected for microvesicles (MVs) isolation and subsequent co-cultured with HT22 cell for sake of assessing their function on neural cell. Relative expression of miR-200c was determined by real time quantitative PCR. Dual luciferase reporter assay was performed to detect the regulatory function of miR-200c to syntaxin-1A. RESULTS: R715 (B1R antagonist) treatment (500 µg/kg) improves neurologic function after cerebral I/R injury indicated by the decrease of Bederson score and infarct volume. MVs from OGD-H/R treated-N9 cell attenuated neural HT22 cell viability, treatment with LDBK (B1R agonist) accelerated the suppression of HT22 resulted from OGD-H/R; whereas this attenuation was partly weakened by B1R antagonist pretreatment (100 nmol/L). At the same time, B1R antagonist pretreatment caused downregulation of miR-200c in N9 cell and N9-derived MVs, and contributed to syntaxin-1A over expression in HT22 cell. Result of luciferase reporter assay suggested that miR-200c can regulate syntaxin-1A expression. MVs from miR-200c knockdown N9 cells medium had the same effect of B1R antagonist that caused the upregulation of syntaxin-1A and improved OGD-H/R-induced reduction of HT22 cell viability. CONCLUSION: Our data suggested that blockage of B1R by B1R antagonist provides neuroprotection action through suppressing signaling delivery of microglia-MVs-miR-200c to neural cell.

Steatosis induced CCL5 contributes to early-stage liver fibrosis in nonalcoholic fatty liver disease progress.

The rapidly increasing prevalence of nonalcoholic fatty liver disease (NAFLD) has become one of the major public health threats in China and worldwide. However, during the development of NAFLD, the key mechanism underlying the progression of related fibrosis remains unclear, which greatly impedes the development of optimal NAFLD therapy. In the current study, we were endeavored to characterize a proinflammatory cytokine, CCL5, as a major contributor for fibrosis in NAFLD. The results showed that CCL5 was highly expressed in fatty liver and NASH patients. In NAFLD rats induced by 8-week-HFD, CCL5 and its receptor, CCR5, were significantly up-regulated and liver fibrosis exclusively occurred in this group. In addition, we showed that hepatocytes are the major source contributing to this CCL5 elevation. Interestingly, a CCL5 inhibitor Met-CCL5, significantly decreased liver fibrosis but not hepatic steatosis. Using a cell model of hepatic steatosis, we found that the conditioned medium of lipid-overloaded hepatocytes (Fa2N-4 cells) which produced excessive CCL5 stimulated the profibrotic activities of hepatic stellate cells (LX-2) as manifested by increased migration rate, proliferation and collagen production of LX-2 cells. CCL5 knockdown in Fa2N-4 cells, Met-CCL5 or CCR5 antibody treatment on LX-2 cells all significantly inhibited the conditioned medium of FFA-treated Fa2N-4 cells to exert stimulatory effects on LX-2 cells. Consistently, the conditioned medium of Fa2N-4 cells with CCL5 over-expression significantly enhanced migration rate, cell proliferation and collagen production of LX-2 cells. All these results support that CCL5 produced by steatotic hepatocytes plays an essential role in fibrotic signaling machinery of NAFLD. In addition, we were able to identify C/EBP-ß as the up-stream regulator of CCL5 gene transcription in hepatocytes treated with free fatty acid (FFA). Our data strongly supported that CCL5 plays a pivotal regulatory role in hepatic fibrosis during NAFLD, which constitutes a novel and exciting observation that may call for potential future development of specific CCL5-targeted NAFLD therapy.

Correlation of hippocampal volume and cognitive performances in patients with either mild cognitive impairment or Alzheimer's disease.

OBJECTIVE: To evaluate hippocampal volume changes and neuropsychological performances in patients with either amnestic mild cognitive impairment (aMCI) or Alzheimer's disease (AD). METHODS: Thirty-eight AD dementia, 22 aMCI patients, and 20 healthy controls were enrolled. Bilateral hippocampal volume was measured concurrently with mini-mental state examination (MMSE), auditory verbal learning test (AVLT), Boston naming test (BNT), and activities of daily living (ADL) test. Baseline and two additional follow-up examinations were conducted. RESULTS: Baseline hippocampal volumes were significantly smaller in AD group than that in aMCI and control groups. MMSE, AVLT, ADL, and BNT scores for the AD group were significantly different from that of both aMCI and control groups. Baseline hippocampal volumes were positively correlated with MMSE and AVLT scores in AD and aMCI patients. At follow-up, left hippocampal volume loss was positively correlated with decreased MMSE and AVLT scores both in AD and aMCI groups, while right hippocampal volume loss was positively associated with decreased AVLT performance only in AD group. Increased ADL and decreased BNT scores were positively associated with left hippocampal volume reduction only in the AD group. CONCLUSIONS: Current findings provide evidence of a close relationship between hippocampal volume and cognitive performances in patients with AD and aMCI, both at baseline and over follow-up.

[Expression profiles and differential diagnostic value of serum Golgi protein-73 in patients with liver cirrhosis and primary hepatic carcinoma].

OBJECTIVE: To investigate the expression profiles of serum Golgi protein-73 (GP73) in liver cirrhosis and primary hepatic carcinoma (PHC) and determine its clinical value for differential diagnosis. METHODS: Serum protein expressions of GP73 and alpha-fetoprotein (AFP) were detected by enzyme-linked immunosorbent assay and chemiluminescence assay, respectively, in patients with PHC (n=80), liver cirrhosis (n=65), and healthy controls (n=50). Inter-group changes were assessed by Kruskal-Wallis test, and significance of these differences was assessed by Mann-Whitney test. A receiver operating characteristic (ROC) curve was plotted to evaluate the diagnostic efficiency and determine the cut-off values for GP73 and AFP. Sensitivity and specificity were compared by the Chi-squared test. Correlation between serum GP73 expression and clinical parameters was determined by Spearman's rank correlation analysis. RESULTS: The PHC group showed significantly higher serum GP73 (282.0 mug/L) than the liver cirrhosis group (211.8 mug/L) and control group (58.3 mug/L) (H = 93.30, P less than 0.01). For differential diagnosis of PHC and liver cirrhosis, the cut-off value was 318.1 mug/L for GP73 and 13.4 mug/L for AFP. Sensitivity of GP73 was lower than AFP (45% (36/80) vs. 65% (52/80); X2 = 8.02, P less than 0.05). Specificity of GP73 was lower than AFP but no significance was found (83.1% (54/65) vs. 87.7% (57/65); X2=0.27, P more than 0.05). The areas under the ROC curves were not significantly different between GP73 and AFP (0.65 (95% confidence interval (CI): 0.54~0.72) vs. 0.75 (95% CI: 0.67~0.83); Z = 1.88, P more than 0.05). The area under the ROC curves increased but not significantly (0.80 (95% CI: 0.73~0.88) vs. 0.75 (95% CI: 0.67~0.83); Z=2.61, P more than 0.05). Serum GP73 was correlated with liver cirrhosis (r=0.27), vascular invasion (r=0.29), and TNM staging (r=0.27) (all P less than 0.05), but not with sex (r=0.13), age (r=0.10), enhanced AFP (> 13.4 mug/L; r=0.03), tumor size (r=0.18), or distant metastasis (r=0.04), all P less than 0.05. CONCLUSION: Serum GP73 and AFP have comparable diagnostic efficiency, but the sensitivity of AFP is superior for differential diagnosis of liver cirrhosis and primary hepatic carcinoma. Elevated serum GP73 may be correlated with liver tumor load and aggressiveness.

[Secretory expression and biological activity analysis of an anti-H5 single-chain antibody from Pichia pastoris].

In our previous study, a panel of 52 broadly cross-reactive H5-specific monoclonal antibodies (MAbs) were generated and characterized. The 13D4, one of these MAbs, has been demonstrated to protect mice against lethal challenge by 4 strains of H5N1 avian influenza virus representing the currently prevailing genetic populations, clades 1, 2.1, 2.2, and 2.3. Here, we further cloned the gene of the 13D4 MAb and constructed a single-chain variable fragment. Then, the 13D4 single-chain antibody (scFv) was expressed in secretory maner in Pichia pastoris. The supernatant of the culture was concentrated and subjected to ammonium sulfate precipitation. The purity of the 13D4 scFv was around 90% in SDS-PAGE following ion-exchange chromatography. We further investigated its binding property using hemagglutination inhibition (HI) test and blocking ELISA. The results indicated that the 13D4 scFv shared the same binding sites and comparable HI titer with the prototype murine 13D4 Mab. In conclusion, an anti-H5 single-chain wide-spectrum neutralizing antibody is prepared successfully in yeast system.

[Analysis on hepatitis C virus genotypes among 118 HCV-infected patients in Xinjiang region].

Not Abstract.