[Association between HBV viral load and severity of liver inflammation in patients with chronic hepatitis B virus infection].

Objective: To explore the relationship and dynamic changes between virological markers and hepatic pathological damage due to host anti-hepatitis B virus (HBV) immunity in the natural course of disease in chronic HBV infected patients. Methods: Two hundred and thirty-eight adult chronic HBV-infected patients who underwent liver biopsy from January 2016 to June 2022 in Taizhou Hospital, Zhejiang Province, were retrospectively selected. General clinical data such as age, gender, platelets, ALT, AST, albumin, HBV DNA, qHBsAg, HBeAg, and liver pathology diagnostic indexes such as the grade of liver necroinflammation and liver fibrotic stages of the patients were collected. The patients were grouped according to HBeAg status, and subgrouped according to different grades of liver necroinflammation and different HBV DNA loads. Statistical analyses were performed to compare the differences in HBV virologic marker levels between the groups, and the correlation between them and the indicators of hepatic inflammatory injury, such as ALT,AST, and the grade of liver necroinflammation in the patients. Results: The levels of HBV virological markers in HBeAg-positive patients with moderate or higher liver necroinflammatory grade (G≥2) were significantly lower than those with mild (no) liver necroinflammatory grade (G < 2) (P < 0.01); whereas the opposite trend was observed in HBeAg-negative patients, with the levels of HBV DNA, and qHBsAg in the G≥2 subgroup being significantly higher than those in the G < 2 subgroup (P < 0.01). Correspondingly, HBV DNA level and qHBsAg showed weak to moderately strong negative correlation with liver necroinflammatory grade and AST which was an indicator of hepatic inflammatory injury in HBeAg-positive patients (P < 0.05); whereas in HBeAg-negative patients, they showed weak to moderately strong positive correlation with hepatic inflammatory activity and ALT, AST (P < 0.001), in which qHBsAg showed only a weak positive correlation with patients' liver necroinflammatory grade (P = 0.003). Further subgroup analyses of HBeAg-positive patients according to whether the HBV DNA level was > 2×10(6) IU/ml showed weak to moderate negative correlations between HBV virological markers and liver necroinflammatory grade as well as ALT and AST in the subgroup of patients with HBV DNA > 2×10(6) IU/ml (P < 0.05); however, the negative correlation disappeared in patients who were still HBeAg positive and had HBV DNA ≤ 2×10(6) IU/ml. Moreover, HBV DNA and ALT, HBeAg and AST showed moderate positive correlation (P < 0.05). Conclusion: We speculate that the activation of host anti-HBV immunity can efficiently inhibit HBV replication by targeting the infected hepatocytes, but only in the early phase of disease progression in HBeAg positive patients with HBV DNA high (> 2×10(6) IU/ml).

[A preliminary discussion on carnosine dipeptidase 1 as a potential novel biomarker for the diagnostic and prognostic evaluation of hepatocellular carcinoma].

Objective: To explore carnosine dipeptidase 1 (CNDP1) potential value as a diagnostic and prognostic evaluator of hepatocellular carcinoma (HCC). Methods: A gene chip and GO analysis were used to screen the candidate marker molecule CNDP1 for HCC diagnosis. 125 cases of HCC cancer tissues, 85 cases of paracancerous tissues, 125 cases of liver cirrhosis tissues, 32 cases of relatively normal liver tissue at the extreme end of hepatic hemangioma, 66 cases from serum samples of HCC, and 82 cases of non-HCC were collected. Real-time fluorescent quantitative PCR, immunohistochemistry, western blot, and enzyme-linked immunosorbent assay were used to detect the differences in mRNA and protein expression levels of CNDP1 in HCC tissue and serum. Receiver operating characteristic (ROC) curves and Kaplan-Meier survival were used to analyze and evaluate the value of CNDP1 in the diagnosis and prognosis of HCC patients. Results: The expression level of CNDP1 was significantly reduced in HCC cancer tissues. The levels of CNDP1 were significantly lower in the cancer tissues and serum of HCC patients than those in liver cirrhosis patients and normal controls. ROC curve analysis showed that the area under the curve of serum CNDP1 in the diagnosis of HCC patients was 0.753 2 (95% CI 0.676-0.830 5), and the sensitivity and specificity were 78.79% and 62.5%, respectively. The combined detection of serum CNDP1 and serum alpha-fetoprotein (AFP) significantly improved the diagnostic accuracy (AUC = 0.820 6, 95% CI 0.753 5-0.887 8). The diagnostic sensitivity and specificity of serum CNDP1 for AFP-negative HCC patients were 73.68% and 68.75% (AUC = 0.793 1, 95% CI 0.708 8-0.877 4), respectively. In addition, the level of serum CNDP1 distinguished small liver cancer (tumor diameter < 3 cm) (AUC = 0.757 1, 95% CI 0.637 4-0.876 8). Kaplan-Meier survival analysis showed that CNDP1 was associated with a poor prognosis in HCC patients. Conclusion: CNDP1 may be a potential biomarker for the diagnostic and prognostic evaluation of HCC, and it has certain complementarity with serum AFP.

IL-17A as a new circulating bioindicator for non-small cell lung cancer diagnosis and prognosis.

OBJECTIVE: Lung cancer (LC) is the highest contributor to cancer-associated mortality worldwide. Approximately 85% of all LC incidences involve non-small cell LC (NSCLC). Unfortunately, owing to a significant lack of sensitive and robust bioindicators, most patient diagnoses occur at advanced stages of the disease, thereby resulting in extremely poor patient outcomes. Herein, we elucidated the role of interleukin-17A (IL-17A) among NSCLC patients. MATERIALS AND METHODS: Circulating IL-17A content was measured using enzyme-linked immunosorbent assay (ELISA), and its diagnostic and prognostic abilities were assessed using the receiver operating characteristic (ROC) curve and Kaplan-Meier analysis, respectively. RESULTS: Our analysis revealed that circulating IL-17A levels were significantly augmented among NSCLC vs. control samples. Moreover, based on our area under the curve (AUC) analysis, circulating IL-17A levels fared considerably better than the standard bioindicator carcinoembryonic antigen (CEA) in both testing and validation cohorts. Notably, we also revealed that the circulating IL-17A levels were accurately and reliably predicted in early-stage NSCLC patients. Besides, we demonstrated a strong correlation between elevated circulating IL-17A expression and worse prognosis among NSCLC patients. CONCLUSIONS: Herein, we demonstrated that circulating IL-17A levels can serve as reliable and potent diagnostic and prognostic bioindicators for NSCLC.

[Natural history and disease progression of chronic hepatitis B virus infection].

OBJECTIVE: To better understand and revise the natural history and disease progression of chronic hepatitis B virus (HBV) infection through analysis of a single-center large-scale cohort of indivi-duals with chronic HBV infection. METHODS: Patients with chronic HBV infection who had undergone liver biopsy in the Fifth Medical Center of Chinese People's Liberation Army (PLA) General Hospital from January 2014 to October 2020 were retrospectively recruited. Based on patient's hepatitis B e antigen (HBeAg) states and pathologic diagnosis, they were categorized into four disease progression statuses (or phases according to the old-terminology in the updated guidelines of chronic hepatitis B (CHB), such as European Association for the Study of the Liver (EASL) 2017, Clinical Practice Guidelines on the Management of Hepatitis B Virus Infection: HBeAg-positive chronic HBV infection (immune tolerance), HBeAg-positive CHB (immune active HBeAg positive), HBeAg-negative chronic HBV infection (inactive carrier), and HBeAg-negative CHB (immune reactive HBeAg negative). Then the demographic, laboratory tests and liver histological results of the patients in different disease progression stages were compared. Age differences between the two groups were evaluated using Mann-Whitney U test. RESULTS: A total of 760 eligible patients with a median age of 29 (interquartile range: 16-39) years were enrolled. Among them, 197 were underage individuals (age < 18 years) and 563 were adults; and 456 were males and 304 females. According to the pathological diagnosis, the patients were classified, and in each of the above four natural disease phases there were 173, 329, 95, and 163 individuals, respectively. Further comparison of the ages of the patients of the four disease progression statuses revealed that patients of HBeAg-negative CHB had a median age at 37 years, which was reasonably higher than those with HBeAg-positive CHB in immune active phase (37 vs. 24 years, P < 0.001), but was relatively younger than those with HBeAg-negative chronic HBV infection (37 vs. 39 years, P= 0.240). CONCLUSION: According to this study, it could be speculated that HBeAg-negative CHB patients probably not all reactivate from individuals of HBeAg-negative chronic HBV infection. Instead, certain HBeAg-negative CHB patients may also come from HBeAg-positive CHB patients who have undergone HBeAg clearance or seroconversion and still remain in the immune active state.

[Consideration on the possible etiological mechanisms and countermeasures about severe acute hepatitis of unknown origin in children].

Since April 2022, severe acute hepatitis of unknown origin in children has spread to 35 countries and regions around the world, and more than 1 010 cases have been reported. Since the severe acute hepatitis of unknown origin involves a wide range of areas and has a high rate, it is critical to identify the etiology and establish effective preventive, diagnostic and therapeutic measures as soon as possible. This study discusses the possible mechanisms and countermeasures of the severe acute hepatitis of unknown origin in children. It speculates that the occurrence of the recent severe acute hepatitis might be related to adenovirus, adeno-associated virus infection, and the COVID-19 epidemic, while the difference in HLA polymorphism among different races might be related to the fact that reported cases were more common in Europe and the United States. Based on the currently available evidence, it can be preliminarily judged that the risk of large-scale outbreak of severe acute hepatitis of unknown origin in children would be low in China, but the persistent awareness and vigilance of the etiology is still needed.

[Clinical application of serum Golgi protein 73 in patients with chronic liver diseases].

Golgi protein 73 (GP73) is a transmembrane protein on the Golgi apparatus and can be cut and released into the blood. In recent years, an increasing number of clinical studies have shown that the elevated serum GP73 level is closely related to liver diseases. And thus GP73 is expected to be used as a new serum marker for assessing progress of chronic liver diseases. Herein, the clinical application of serum GP73 in chronic hepatitis, liver fibrosis, liver cirrhosis and hepatocellular carcinoma with different etiologies was reviewed based on available literatures; and a research outlook in this field is made.

[Diagnostic value of liver stiffness measurement for the evaluation of liver fibrosis in nonalcoholic fatty liver].

Objective: To investigate the clinical and diagnostic value of liver stiffness measurement (LSM) for the evaluation and comparison of aspartate aminotransferas/platelet ratio index (APRI), fibrosis 4 indexes (FIB-4) and NAFLD fibrosis score (NFS) with liver fibrosis staging in relation to nonalcoholic fatty liver disease (NAFLD). Methods: 103 cases with NAFLD who met the inclusion criteria confirmed by liver biopsy were selected for retrospective analysis. The results of serological tests and LSM were recorded. The APRI, FIB-4 and NFS were calculated. The accuracy and applicability of four liver fibrosis models in the diagnosis of liver fibrosis in NAFLD patients were compared with the receiver operating characteristic curve (ROC), and the diagnostic cut-off value of LSM was established. Results: Varying degrees of LSM, APRI, FIB-4 and NFS had shown positive correlations with the increasing degree of liver fibrosis. Among them, LSM was positively correlated with the degree of liver fibrosis, and the correlation coefficient was r = 0.727, P < 0.0001. Consistent with this, the area under the receiver operating characteristic curve, sensitivity, and specificity of LSM diagnosis of liver fibrosis in different stages was significantly higher than APRI, FIB-4 and NFS. Area under receiver operating characteristic curve of LSM was 0.862 and 0.928 for significant liver fibrosis (f ≥ 2), and advanced liver fibrosis (f ≥ 3). Conclusion: LSM has a good diagnostic exclusion value for NAFLD-induced fibrosis, and its sensitivity and specificity are better than APRI, FIB-4 and NFS.

[Establishment and preliminary application of serum Golgi protein 73 based noninvasive diagnostic model for compensated stage hepatitis B cirrhosis].

Objective: To establish and evaluate diagnostic efficacy and applicability of serum Golgi protein (GP) 73 based non-invasive diagnostic model with other conventional serological indicators for compensated stage hepatitis B cirrhosis. Methods: 666 cases with chronic hepatitis B (CHB) who had visited to the Fifth Medical Center of People's Liberation Army General Hospital from January 2010 to December 2017 were selected as the study subjects, and were classified according to compensated stage cirrhosis into clinical and pathological diagnosis group based on whether or not the liver histological examination was performed. A diagnostic model of compensated stage hepatitis B cirrhosis in the clinical diagnosis group was established. The current clinically used diagnostic model of liver cirrhosis, aspartate aminotransferase/platelet ratio index (APRI), fibrosis index (FIB)-4 and liver stiffness measurement (LSM) were compared. Eventually, the diagnostic model was verified step by step by pathological diagnosis group. Results: The area under the receiver operating characteristic curve (AUC) of GP73 and APRI, FIB-4, and LSM for cirrhosis patients in the clinical diagnosis group were 0.842, 0.857, 0.864, and 0.832, respectively. The diagnostic efficiency of the four indicators were of similar (P value > 0.05). A diagnostic model of compensated stage hepatitis B cirrhosis (GAPA) using logistic regression analysis was established: LogitP = 1/ [1 + exp (1.614-0.054 × GP73-0.045 × Age + 0.030 × PLT-0.015 × ALP)]. The AUC of the model was as high as 0.940 and the optimal cut-off value were 0.41. The corresponding diagnostic sensitivity and specificity were 0.92 and 0.82, respectively. The diagnostic efficiency was better than that of APRI, FIB-4, LSM and GP73 alone (P < 0.05). The AUC of GAPA was 0.877 in the pathological diagnosis group, which was similar to the diagnostic efficacy of LSM (0.891) and FIB-4 (0.847) (P > 0.1), but still superior to that of APRI (0.811) and GP73 alone (0.780) (P < 0.001). Conclusion: GAPA, a diagnostic model for compensated stage hepatitis B cirrhosis established in this study, has a good diagnostic efficacy in both the clinical and pathological diagnosis group, and has certain auxiliary diagnostic value in the areas where resources are relatively scarce or where LSM has not been developed.

[Expression and clinical significance of chemokine CXCL10 and its receptor CXCR3 in hepatocellular carcinoma].

OBJECTIVE: To explore the expression and clinical significance of chemokine CXCL10 and CXCR3 in hepatocellular carcinoma (HCC). METHODS: The expression and prognostic of CXCL10 and CXCR3 in HCC tumor tissues and non-tumor tissues were analyzed in two different publicly available databases the Cancer Genome Atlas (TCGA) and Liver Cancer Institute (LCI). In addition, quantitative real-time PCR (qPCR) was used to detect the mRNA expression of CXCL10 and CXCR3 in 45 HCC clinical samples with HBV infection background. Pearson correlation and Spearman rank correlation were used to determine the correlation between the expression level of CXCL10 and CXCR3 in tumor and non-tumor tissues. RESULTS: In TCGA database, the expression of CXCL10 in HCC tumor tissues was significantly higher than that in non-tumor tissues (nonpaired samples: 3.379±2.081 vs. 2.213±2.274, P<0.001; paired samples: 3.159±2.267 vs. 2.213±2.274, P=0.018). Similarly in LCI datebase (7.625±1.683 vs. 7.287±1.328, P=0.009). And higher CXCL10 expression was significantly associated with a better prognosis in the patients with HCC both in TCGA and LCI database (P=0.107, P=0.002). In TCGA database, the expression of CXCR3 in HCC tumor tissues was significantly higher than that in non-tumor tissues (nonpaired samples: -0.906±1.697 vs. -1.978±1.629, P<0.001; paired samples: -1.329±1.732 vs. -1.978±1.629, P=0.037), while lower in LCI database (3.989±0.339 vs. 4.074±0.309, P=0.003). In both databases, higher CXCR3 expression was significantly associated with a better prognosis in the HCC patients (P=0.004, P=0.014). Furthermore, in TCGA database, the expression level of CXCL10 and CXCR3 was positively correlated both in HCC tumor tissues and matched non-tumor tissues (r=0.584, P<0.001; r=0.776, P<0.001). The qPCR assay showed that the expression of CXCL10 in HBV-related HCC tumor tissues was significantly higher than those in normal liver tissues [0.479(0.223, 1.094) vs. 0.131(0.106, 0.159), P=0.010], and the expression in HBV-related non-tumor tissues was also significantly higher than those in normal liver tissues [0.484(0.241, 0.846) vs. 0.131(0.106, 0.159), P<0.001]. The same was true as CXCR3 [0.011(0.006, 0.019) vs. 0.002(0.001, 0.004), P=0.004; 0.016(0.011, 0.021) vs. 0.002(0.001, 0.004), P<0.001]. However there was no significant difference of CXCL10 and CXCR3 between tumor tissues and matched non-tumor tissues (P=1.000, P=0.374). CONCLUSION: Expression of CXCL10 was up-regulated in HCC tissues, expression of CXCR3 was down-regulated in HBV-related HCC tissues, and the higher expression of both genes was correlated with better overall survival in HCC patients.

[Re-recognition of occult hepatitis B virus infection].

Hepatitis B virus (HBV) infection remains to be a serious public health problem in China. There used to be a high prevalence of HBV infection in China, which resulted in a large number of HBV susceptible and post-infected population. Single anti-HBc positive usually indicates post HBV infection and its prevalence is particularly high among people over 40 years old, some of whom may be occult hepatitis B virus infection (OBI). The clinical diagnosis of OBI is difficult and easily missed. Since OBI may cause chronic liver disease progression and even lead to cirrhosis and hepatocellular carcinoma eventually, and more importantly, patients with OBI may leed to HBV reactivation when the immune function decreases or immunosuppressive therapy is performed, the accurate identify of OBI is of particular importance. Moreover, OBI is the potential source of HBV infection, which may transmit through blood transfusion, organ transplantation and mother-to-child transmission. In view of this situation, we reviewed the mechanism, prevalence and definition of OBI, and proposed a determination system for replication-competent HBV DNA based on our understanding of the updated OBI definition. It is expected to be beneficial for OBI diagnosis, treatment and management.

[Mechanism relevant to hepatocellular carcinoma occurrence after negative conversion of viral DNA in treatment of chronic hepatitis B patients with nucleos(t)ide drugs].

Hepatocellular carcinoma (HCC) is a common malignant tumor in China, and most of the patients have a background of chronic HBV infection. Nucleos(t)ide drugs (NAs) are currently recommended by major guidelines as a first-line treatments for chronic hepatitis B. However, it is still clinically possible to observe that some patients who have acquired virological response (HBV DNA below the lower detection limit) after NAS treatment progress to HCC, and its mechanism of development is still unclear. In this review, the mechanism relevant to HCC progression in treatment of chronic hepatitis B patients with NAs is analyzed mainly from the aspects of gene integration and persistent inflammatory injury.

[Prognostic significance of albumin/globulin ratio on postoperative survival of patients with hepatocellular carcinoma].

Objective: To investigate the prognostic value of albumin/globulin ratio on postoperative survival outcomes in patients with hepatocellular carcinoma. Methods: Data of 630 patients with HCC, who underwent surgical resection from February 2009 to July 2013, were retrospectively analyzed. Patients were divided into low-value group (A/G < 1.5, defined as L group) and high-value group (A/G≥1.5, defined as H group), and their distribution characteristics were observed with the normal A/G threshold value. Independent risk factors' affecting survival and prognosis was analyzed with univariate and multivariate Cox's regression model. Survival trend of all patients with low-value and high-value groups in A, B and C of Barcelona stage (BCLC stage) were analyzed using the Kaplan-Meier method. Results: Multivariate analysis showed that preoperative A/G ratio (P = 0.007), alpha-fetoprotein (P < 0.001), gamma-glutamyltransferase (P = 0.006), RBC (P = 0.014), international normalized ratio (P = 0.009), preoperative BCLC staging (P < 0.001) and number of tumors (P = 0.003), and intraoperative blood transfusion (P < 0.001) were independent prognostic factors affecting long-term survival in HCC patients. The median overall survival time in-group L was 15 months, significantly lower than that in group H of 42 months (P < 0.001). Stratified analysis showed that the short-term survival advantage of patients with high A / G value was limited to those with Barcelona stage A (P < 0.001), and disappeared in patients with Barcelona stage B and C (P > 0.05). The long-term survival advantage existed in patients with Barcelona stage A (P < 0.001), B (P < 0.05), and disappeared in C (P > 0.05). Conclusion: Preoperative albumin/globulin ratio can predict postoperative prognosis and survival, and direct towards the treatment for early stage of HCC and thus representing as an indicator of high clinical value.

[Diagnostic application of serum GP73 and the relevant mechanism in the diagnosis of liver cirrhosis].

Liver cirrhosis is the end-stage change of chronic liver diseases with various causative factors. The accurate diagnosis of liver cirrhosis in an early stage is very important for the timely treatment and prognosis of patients. A liver biopsy test is the gold standard for the diagnosis of liver cirrhosis; but its use is limited in clinical practices due to its invasive nature. The conventional non-invasive measures (APRI, FIB-4 and LSM) for diagnosis of liver cirrhosis could not fulfill the needs. Therefore, finding a new serological marker of liver cirrhosis has become a research hotspot. Based on literature review and our own results, we suggest that serum GP73 as the most potential serum marker for liver cirrhosis diagnosis. This review briefly introduces the application of serum GP73 in the diagnosis of liver cirrhosis and the potential mechanism relevant to its involvement in the development of liver cirrhosis.

[Hepatitis B virus X protein-regulated expression of Plk1].

OBJECTIVE: To investigate the ability and underlying mechanism of hepatitis B virus X protein (HBx) regulation of Polo-like kinase 1 (Plk1) expression. METHODS: The human HCC cell line HepG2 was transfected (transiently and stably) with an HBx plasmid expression vector (pCMV-HA-HBx) or empty plasmid vector (control), with and without expression plasmids with the Plk1 promoter. Effects on Plk1 expression were assessed by western blotting. Functional effects on the Plk1 promoter were assessed by luciferase reporter assay. Effects on the mRNA level of Plk1 in S phase HepG2 cells were assessed by quantitative real-time reverse transcriptase polymerase chain reaction. After blocking protein synthesis by treatment with cycloheximide (CHX), the turnover rate of Plk1 was assessed by western blotting. Lastly, the effect of HBx on cell cycle was assessed by flow cytometry. RESULTS: HBx did not increase the protein expression of Plk1 in non-synchronized HepG2 cells, but did significantly up-regulate the Plk1 protein level in the synchronized S phase cells (P = 0.026 and P = 0.003, respectively). Ectopic expression of HBx did not increase the mRNA level of Plk1 in HepG2 cells, but did inhibit the degradation of Plk1, as evidenced by an increased half-life of Plk1 protein (from 30 to 90 minutes). The HBx-expressing HepG2 cells showed more frequent entry into the S or G(2)/M phase than the control cells (31.65% vs. 24.56% or 9.43% vs. 4.47%, respectively) and less in the G(0)/G(1) phase (decrease from 70.97% to 58.92% for the HBx-expressing HepG2 cells). CONCLUSION: HBx is able to up-regulate the expression of Plk1 in HepG2 cells by a mechanism involving stabilization of the Plk1 protein primarily in the S phase of the cell cycl.

Effect of functional nuclear factor-kappaB genetic polymorphisms on hepatitis B virus persistence and their interactions with viral mutations on the risk of hepatocellular carcinoma.

BACKGROUND: Nonresolving inflammation and viral mutations are important in hepatitis B virus (HBV)-induced hepatocarcinogenesis. However, the effects of genetic polymorphisms affecting nuclear factor-kappaB (NF-κB) on HBV persistence and generation of hepatocellular carcinoma (HCC)-related HBV mutations remain unknown. PATIENTS AND METHODS: rs28362491 (NFKB1 -94Ins > Del), rs2233406 (NFKBIA -826C > T), rs3138053 (NFKBIA -881A > G), and rs696 (NFKBIA +2758G > A) were genotyped in 1342 healthy controls, 327 HBV-clearance subjects, and 3976 HBV-positive subjects including 1495 HCC patients, using quantitative PCR. HBV mutations were determined by sequencing. The NFKBIA promoter activity was assessed by transient transfection. Multiplicative interactions of the polymorphisms and viral mutations were assessed by multivariate logistic regression. RESULTS: Compared with HBV-clearance subjects, rs2233406 (CT versus CC) and rs3138053 (AG or AG + GG versus AA) significantly decreased HBV persistence, especially in the genotype B HBV-infected subjects. In the genotype C HBV-infected subjects, rs2233406 variant genotypes were significantly associated with an increased risk of HCC [CT versus CC: age-, gender-adjusted odds ratio (AOR), 1.33; 95% confidence interval (CI) 1.01-1.75 in training set and AOR, 1.59; 95% CI 1.01-2.52 in validation set] compared with HCC-free HBV-infected subjects and significantly increased the frequencies of HCC-related HBV mutations (A1762T/G1764A, T1753V, preS1 start codon mutation, and preS deletion); rs28362491 (Del/Del or Ins/Del + Del/Del versus Ins/Ins) significantly increased the frequency of A1762T/G1764A and reduced the frequency of preS2 start codon mutation. The variant genotypes impaired NFKBIA promoter activity in hepatic cells. The interaction of rs2233406 variant genotypes (CT + TT versus CC) with A1762T/G1764A significantly increased HCC risk in genotype C HBV-infected subjects, with AOR of 2.61 (95% CI 1.09-6.26). CONCLUSION: Genetic polymorphisms improving NF-κB activity contribute to genotype B HBV clearance. The rs2233406 variant genotypes significantly increase HCC risk, possibly via facilitating immune selection of the HBV mutations. The host-virus interactions are important in identifying HBV-infected subjects who are more likely to develop HCC.

Unique features of PTCH1 mutation spectrum in Chinese sporadic basal cell carcinoma.

BACKGROUND: Alterations of the PTCH1 gene have been found to contribute to both familial and sporadic basal cell carcinoma (BCC), especially in Caucasian patients. Furthermore, the majority of PTCH1 gene mutations in sporadic BCCs in Caucasian patients carry ultraviolet (UV) signatures, suggesting the key role of UV light in BCC development. However, sporadic BCC in non-Caucasian population has a lower incidence, and the pathogenesis remains largely unknown. To date, there has been no mutation analysis on PTCH1 gene in Chinese patients with sporadic BCCs. OBJECTIVE: To investigate genetic alterations of the PTCH1 gene in Chinese sporadic BCCs. METHODS: Direct sequencing was used to screen for mutations in PTCH1 in 31 microdissected samples in Chinese sporadic BCCs. In addition, single nucleotide polymorphisms (SNPs) were studied for loss of heterozygosity (LOH). RESULTS: Nineteen PTCH1 mutations in 17 of the 31 BCCs (54.8%) were identified. SNP analysis revealed LOH of PTCH1 in 10 of 23 BCCs (43.5%). Interestingly, the majority of mutations identified (63.2%) were insertion/deletion, which was different from the results in Caucasian cases whose mutations are predominantly point mutations. Only two (10.5%) of the remaining seven mutations were UV-specific C → T transition or tandem CC → TT transitions. All mutations occurred evenly throughout the entire PTCH1 protein domain without a hot-spot detected. CONCLUSION: Mutations and LOH in PTCH1 were also highly prevalent in Chinese sporadic BCCs. However, UV light plays a less role in causing these mutations, suggesting other potential mechanisms in the development of sporadic BCC in Chinese patients.

Induction treatment of proliferative lupus nephritis with leflunomide combined with prednisone: a prospective multi-centre observational study.

To evaluate the efficacy and safety of leflunomide in the treatment of proliferative lupus nephritis, a prospective multi-centre observational study was conducted. Patients with biopsy proven proliferative lupus nephritis were assigned to receive either leflunomide or cyclophosphamide with concomitant prednisone. Leflunomide was given orally with a loading dose of 1 mg/kg/day for 3 days followed by 30 mg/day. Intravenous cyclophosphamide was administered monthly at a dose of 0.5 g/m2 of body-surface area. A total of 110 patients were enrolled, 70 in the leflunomide group and 40 in the cyclophosphamide group. The complete remission rate in the leflunomide group was 21% and partial remission rate 52%, as compared with 18% and 55%, respectively, in the cyclophosphamide group. Renal parameters and systemic lupus erythematosus disease activity index improved significantly and similarly in both groups. Serum creatinine decreased or stabilized in both treatment groups. No significant difference was noted with respect to clinical outcome between groups. Repeat biopsy also showed a significant reduction of active lesions in kidney pathology after 6 months of leflunomide treatment. Major adverse events, similar in both treatment groups, included infection, alopecia and hypertension. Leflunomide, compared with cyclophosphamide, in combination with prednisone was effective in the induction therapy of proliferative lupus nephritis and was generally well-tolerated.

Mutation analysis of Jagged1 (JAG1) in Alagille syndrome patients.

Alagille syndrome (AGS) is an autosomal dominant disorder caused by mutations in Jagged1 (JAG1), a ligand in the evolutionarily conserved Notch signaling pathway. Previous studies have demonstrated that a wide spectrum of JAG1 mutations result in AGS. These include total gene deletions, protein truncating, splicing and missense mutations which are distributed across the coding region of the gene. Here we present results of JAG1 mutation screening by SSCP and FISH in 105 patients with AGS. For these studies, new primers were designed for 12 exons. Mutations were identified in 63/105 patients (60%). The spectrum of the JAG1 mutations presented here is consistent with previously reported results. Eighty three percent (52/63) of the mutations were protein truncating, 11% (7/63) were missense, 2% (1/63) were splice site, and 5% (3/63) were total gene deletions demonstrable by FISH. Six of the missense mutations are novel. As has been reported previously, there is no apparent relationship between genotype and clinical phenotype.

Two-chain high molecular weight kininogen induces endothelial cell apoptosis and inhibits angiogenesis: partial activity within domain 5.

We previously reported that the binding of two-chain high molecular weight kininogen (HKa) to endothelial cells may occur through interactions with endothelial urokinase receptors. Since the binding of urokinase to urokinase receptors activates signaling responses and may stimulate mitogenesis, we assessed the effect of HKa binding on endothelial cell proliferation. Unexpectedly, HKa inhibited proliferation in response to several growth factors, with 50% inhibition caused by approximately 10 nM HKa. This activity was Zn(2+) dependent and not shared by either single-chain high molecular weight kininogen (HK) or low molecular weight kininogen. HKa selectively inhibited the proliferation of human umbilical vein and dermal microvascular endothelial cells, but did not affect that of umbilical vein or human aortic smooth muscle cells, trophoblasts, fibroblasts, or carcinoma cells. Inhibition of endothelial proliferation by HKa was associated with endothelial cell apoptosis and unaffected by antibodies that block the binding of HK or HKa to any of their known endothelial receptors. Recombinant HK domain 5 displayed activity similar to that of HKa. In vivo, HKa inhibited neovascularization of subcutaneously implanted Matrigel plugs, as well as rat corneal angiogenesis. These results demonstrate that HKa is a novel inhibitor of angiogenesis, whose activity is dependent on the unique conformation of the two-chain molecule.

The activation of protein kinase C induces higher production of reactive oxygen species by mononuclear cells in patients with multiple sclerosis than in controls.

OBJECTIVE: Recent findings have increasingly shown the importance of reactive oxygen species (ROS) in causing oxidative damage to macromolecules and in contributing to tissue degeneration in target organs of autoimmune diseases. This study was aimed at comparing the base line and induced production of ROS by peripheral blood mononuclear cells (PB MNCs) of patients with multiple sclerosis (MS) in remission and relapse, of patients with other neurological diseases (OND) and of healthy controls. In addition, we analyzed the underlying mechanism of ROS production. METHODS: PB MNCs were separated from 28 MS patients in remission and 13 in relapse, and from 29 healthy controls and 10 OND. ROS was measured by spectrofluorometry. Expression of proinflammatory cytokines was assessed by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Mitochondrial (mt) DNA haplotypes were determined by using restriction site polymorphism analysis. RESULTS: The base line and tumor necrosis factor (TNF)-alpha or interferon (IFN)-gamma induced ROS values were similar in the four groups, and the individual measures did not show a correlation with MS associated mtDNA haplotypes. Phorbol ester activation of protein kinase C (PKC) induced higher ROS production in all groups, however, with significantly greater values in the MS remission group. Calphostine C, a PKC inhibitor decreased or eliminated ROS production in a dose-dependent manner, suggesting further that it was predominantly or exclusively generated by PKC activated NADPH oxidase. A trend of increased TNF-alpha and IFN-gamma expression was noted in the MS relapse group, in contrast to the high ROS release in the MS remission group. CONCLUSION: The detected phase difference between the highest ROS production vs TNF-alpha expression is compatible with the hypothesis that different subpopulations of monocytes/macrophages are involved. We suggest that the ROS producing subpopulation preferentially migrates into the central nervous system (CNS) during a relapse. The present study together with our previous observation on oxidative damage to DNA in active plaques delineates a molecular pathway likely involved in the histologic evolution of inflammatory demyelination.