[Preliminary evaluation of immunogenicity and protective effect of multicomponent recombinant protein vaccine EPRHP014 against tuberculosis].

Objective: To evaluate the immunogenicity and protective effect of a multicomponent recombinant protein vaccine EPRHP014 constructed independently and provide a scientific basis for developing new tuberculosis (TB) vaccine and effective prevention and control of TB. Methods: Three full-length Mycobacterium (M.) tuberculosis protein antigens (EsxH, Rv2628, and HspX) and two epitope-predicted and optimized epitope-dominant protein antigens (nPPE18 and nPstS1) were selected, from which five protein antigens were used to construct a protein antigen composition EPRHP014, including a fusion expression multi-component protein antigen (EPRHP014f) and a multi-component mixed protein antigen (EPRHP014m) formed with the five single protein using clone, purification, and purification respectively. Multicomponent protein vaccines EPRHP014f and EPRHP014m were prepared with aluminum adjuvant, and the BCG vaccine was used as a control. ELISA detected the titer of serum-specific antibodies, the secretion of various cytokines was detected by ELISpot and Luminex, and immune protection was observed by the M. tuberculosis growth inhibition test in vitro. The results were statistically analyzed by t-test or rank sum test, and P<0.05 was considered a statistically significant difference. Results: Mice Immunized with EPRHP014m and EPRHP014f could produce highly effective IgG antibodies and their subtypes IgG1 and IgG2a, and the antibody titers were similar to those of mice immunized with BCG, with no statistical significance (P>0.05). The number of spot-forming cells (SFC) secreting IFN-γ and IL-4 induced by EPRHP014f group was significantly higher than those by EPRHP014m group and BCG group (P<0.05), but there was no significant difference in the number of SFC for IFN-γ and IL-4 induced between EPRHP014m group and BCG group (P>0.05). The secretion levels of GM-CSF and IL-12p70 induced by the EPRHP014m group were higher than those of the BCG group (P<0.05), but there was no significant difference in the levels of IL-6 and IL-10 induced between EPRHP014m group and BCG group (P>0.05). There was no significant difference in the secretions of IL-6, IL-10, IL-12, and GM-CSF between the EPRHP014f and BCG groups (P>0.05). EPRHP014m group, EPRHP014f group, and BCG group had obvious antibacterial effects in vitro, and the difference was insignificant (P>0.05). Conclusion: Both EPRHP014f and EPRHP014m can induce strong humoral and cellular immune responses in mice after immunization, and have a strong ability to inhibit the growth of M. tuberculosis in vitro, indicating that the antigen composition EPRHP014 has good potential in the development and application of TB vaccine.

Unintrusive multi-cancer detection by circulating cell-free DNA methylation sequencing (THUNDER): development and independent validation studies.

BACKGROUND: Early detection of cancer offers the opportunity to identify candidates when curative treatments are achievable. The THUNDER study (THe UNintrusive Detection of EaRly-stage cancers, NCT04820868) aimed to evaluate the performance of enhanced linear-splinter amplification sequencing, a previously described cell-free DNA (cfDNA) methylation-based technology, in the early detection and localization of six types of cancers in the colorectum, esophagus, liver, lung, ovary, and pancreas. PATIENTS AND METHODS: A customized panel of 161 984 CpG sites was constructed and validated by public and in-house (cancer: n = 249; non-cancer: n = 288) methylome data, respectively. The cfDNA samples from 1693 participants (cancer: n = 735; non-cancer: n = 958) were retrospectively collected to train and validate two multi-cancer detection blood test (MCDBT-1/2) models for different clinical scenarios. The models were validated on a prospective and independent cohort of age-matched 1010 participants (cancer: n = 505; non-cancer: n = 505). Simulation using the cancer incidence in China was applied to infer stage shift and survival benefits to demonstrate the potential utility of the models in the real world. RESULTS: MCDBT-1 yielded a sensitivity of 69.1% (64.8%-73.3%), a specificity of 98.9% (97.6%-99.7%), and tissue origin accuracy of 83.2% (78.7%-87.1%) in the independent validation set. For early-stage (I-III) patients, the sensitivity of MCDBT-1 was 59.8% (54.4%-65.0%). In the real-world simulation, MCDBT-1 achieved a sensitivity of 70.6% in detecting the six cancers, thus decreasing late-stage incidence by 38.7%-46.4%, and increasing 5-year survival rate by 33.1%-40.4%, respectively. In parallel, MCDBT-2 was generated at a slightly low specificity of 95.1% (92.8%-96.9%) but a higher sensitivity of 75.1% (71.9%-79.8%) than MCDBT-1 for populations at relatively high risk of cancers, and also had ideal performance. CONCLUSION: In this large-scale clinical validation study, MCDBT-1/2 models showed high sensitivity, specificity, and accuracy of predicted origin in detecting six types of cancers.

[Study of the inhibitory effect of cell entry inhibitory ezetimibe on hepatitis B virus in vitro].

Objective: To study the inhibitory effect of ezetimibe in an experimental model of human hepatoma cell line (HepaRG) infected with hepatitis B virus (HBV) positive human serum in vitro. Methods: Mature HepaRG cells were divided into a treatment group (received drugs) and a control group (did not receive drugs). In the ezetimibe prevention experiment, the cells in the treatment group was treated with drugs 2 h before infection and 24 h during infection. In the ezetimibe treatment experiment, the cells in the treatment group were treated with drugs for 6 ~ 10 days continuously after 24 hours of HBV infection. The expression of HBV DNA and intracellular cccDNA in the supernatant was detected by fluorescence quantitative PCR. Hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) content in the cell supernatant were detected by chemiluminescence. Analysis of variance was used to compare the differences between multiple groups. Pairwise comparisons among groups were followed by t- test with normal distribution. P < 0.05 was considered as statistically significant. Results: Ezetimibe prevention experiment showed that compared with control group, the treatment group was added with 20, 60, and 100 µmol/L ezetimibe before and during infection, and the HBV DNA content in the supernatant 2 days before was significantly reduced (P < 0.05) in the treatment group. Compared with the control group, the HBsAg expression level 2 days before was significantly reduced (P < 0.05) with the addition of 60 µmol/L ezetimibe in the treatment group. Compared with the control group, the expression level of intracellular cccDNA was significantly reduced (P < 0.05) after 10 days with the addition of 100µmol/L ezetimibe in the treatment group. Ezetimibe treatment experiment showed that cccDNA content in the cells were significantly lowered with the immediate addition of 60µmol/L ezetimibe 24 hours after infection for 10 days when compared to control group (P < 0.05). Conclusion: Ezetimibe, as a cytosolic inhibitor, has a certain inhibitory effect on hepatitis B virus infection in both prevention and treatment experiment.

[A new exploration and consideration of Type A fracture of distal femur based on finite element biomechanical analysis].

Objective: To investigate the changes of internal fixation stress under different angles of interior fracture line and different screw placement modes in the case of A-type distal femoral fracture. Methods: A 24-year-old healthy male volunteer was recruited to collect the right femur data. CATIA V5R21 software produced a 10 mm fracture gap at the external side of the femur 6.5 cm proximal to the joint line and different angle fracture lines were generated on the internal of the femur at the same height. Based on the actual measured dimensions, the three-dimensional (3D) model of the locking plate and screw was reconstructed using CATIA V5R21 software, ignoring the screw surface threads and then the assembly of the internal fixation of the titanium plate, screws and femur was done. All models were meshed using Hypermesh 13.0 software. The assembled 3D model was input into ABAQUS 6.14 to generate a finite element model. Preliminary finite element biomechanical analysis was performed using the four medial fracture line angles and the stress distribution of the internal fixation under the three screw placement modes, and then the analysis was continued after the optimal screw placement method was re-determined. Results: Under an axial loading of 700 N, with the increase of the angle of the fracture line, the stress of the lateral internal fixation gradually increased, and the displacement of the proximal end of the fracture gradually increased. The sequential screw placement method was superior to the leaping screw placement method. The placement of the first screw at the proximal end of the fracture was critical to the distribution of the internal fixation stress. Conclusions: The operation plan of the type A of distal femoral fracture needs to be confirmed according to the internal and external fracture's condition. When the fracture line is at a excessive positive angle or a negative angle, a simple lateral fixation may not provide a stable fracture fixation so that other fixation methods are needed.

MicroRNA-15a-5p down-regulation inhibits cervical cancer by targeting TP53INP1 in vitro.

OBJECTIVE: An increasing number of reports have shown that microRNAs (miRNAs) play a vital role in the occurrence and development of cancer by acting as tumor inhibitors or oncogenes. The purpose of this research was to explore whether the expression level of microRNA-15a-5p (miR-15a-5p) was related to TP53 regulated inhibitor of apoptosis 1 (TP53INP1) in cervical cancer, and to explore the role of miR-15a-5p in cervical cancer in vitro. PATIENTS AND METHODS: Human cervical cancer tissues and adjacent normal tissues were obtained from 30 cervical cancer patients. Firstly, we carried out the quantitative Real Time-PCR (qRT-PCR) assay to evaluate the level of miR-15a-5p in cervical cancer tissues and cell lines. The TargetScan and the Dual-Luciferase Reporter Assay were used to confirm the relationship between TP53INP1 and miR-15a-5p. Besides, the Cell Counting Kit-8 (CCK-8) and the flow cytometry analysis were performed to detect the effect of miR-15a-5p on cell proliferation and apoptosis in cervical cancer cells. RESULTS: Our results showed that the expression of miR-15a-5p was enhanced in cervical cancer tissues and cells lines. The data from the Dual-Luciferase Reporter Assay demonstrated that TP53INP1 was a direct target of miR-15a-5p. We also found that TP53INP1 was down-regulated in the cervical cancer tissues and cell lines compared with the adjacent normal tissues and normal cervical cells. Besides, the down-regulation of miR-15a-5p depressed cervical cancer cell proliferation and enhanced cell apoptosis. Our results clearly suggested that the down-regulation of TP53INP1 successfully impaired the tumor-inhibition effects of miR-15a-5p inhibitor in cervical cancer cells. CONCLUSIONS: Our findings indicated that miR-15a-5p functioned as a tumor-promoting gene in cervical cancer by directly targeting TP53INP1, indicating that miR-15a-5p might be a potential treatment target for cervical cancer patients.

MicroRNA-449b-5p promotes the progression of osteoporosis by inhibiting osteogenic differentiation of BMSCs via targeting Satb2.

OBJECTIVE: We aimed to explore the role of microRNA-449b-5p in osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and its mechanism of action. MATERIALS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) assay was used to detect the expression levels of microRNA-449b-5p and osteogenic markers including RUNX2, OCN during BMSCs differentiation. The microRNA-449b-5p mimic and microRNA-449b-5p inhibitors were transfected into BMSCs to achieve microRNA-449b-5p overexpression and knockdown, then the expressions of osteogenic markers were detected by qRT-PCR. The ALP activity staining and the alizarin red staining were used to detect the activity of ALP and the mineralization ability of cells after overexpression and knockdown of microRNA-449b-5p. Binding sites for microRNA-449b-5p and Satb2 were predicted by TargetScan, the PicTar and microRNAanda programs, and confirmed by dual-luciferase reporter gene assay. The relationship between microRNA-449b-5p and Satb2 was analyzed by QRT-PCR and Western blot. The microRNA-449b-5p inhibitor and shSATB2 lentivirus were simultaneously transfected in BMSCs, and the expression levels of RUNX2, OCN and ALP were detected by qRT-PCR and ALP activity assays. RESULTS: microRNA-449b-5p expression gradually decreased during osteogenic differentiation. Overexpression of microRNA-449b-5p inhibited BMSCs differentiation by down-regulating ALP activity, RUNX2, and OCN expression, while the opposite result was observed after knockdown of microRNA-449b-5p. MicroRNA-449b-5p can bind to the 3'UTR end of Satb2, which was involved in the osteogenic differentiation of microRNA-449b-5p-regulated BMSCs, and silencing of Satb2 can abolish the positive effect of the microRNA-449b-5p inhibitor on osteoblasts differentiation. CONCLUSIONS: microRNA-449b-5p could aggravate osteoporosis by inhibiting osteogenic differentiation of BMSCs through targeting Satb2.

[The progress and prospect of ophthalmological research on cerebral visual impairment in children].

With the rapid development of global perinatal medicine and neonatal medicine, the survival rate of high-risk neonates (premature, and those who suffer neurologic lesion during or shortly after birth, et al.) improved significantly, however the incidence of cerebral visual impairment (CVI) also rose, which has become the most common cause of visual impairment in children in developed countries. Studies found that visual abnormalities of patients with CVI can be various. Since children's cognition and motor development are inseparable from visual functions, children with CVI are usually characterized by abnormalities in sensory perception, cognition and even movement, other than visual impairment itself. Due to the characteristics of CVI, such as early onset, complex etiology, difficulty in diagnosis and treatment, and involvement with ophthalmology, pediatrics, rehabilitation medicine, genetic epidemiology and other multi-disciplinary content, current domestic research on CVI is limited. From the perspective of ophthalmologist, this paper reviews the progress of ophthalmology diagnosis and treatment in children with CVI in recent years, aiming to have better early recognition and individualized intervention, so as to help pediatrician and rehabilitation physicians to improve survival skills for CVI children and their quality of life. (Chin J Ophthalmol, 2019, 55:469-474).

[Pilot study of the relationship between clinical classification of gallbladder cancer and prognosis: a retrospective multicenter clinical study].

Objectives: To propose a novel clinical classification system of gallbladder cancer, and to investigate the differences of clinicopathological characteristics and prognosis based on patients who underwent radical resection with different types of gallbladder cancer. Methods: The clinical data of 1 059 patients with gallbladder cancer underwent radical resection in 12 institutions in China from January 2013 to December 2017 were retrospectively collected and analyzed.There were 389 males and 670 females, aged (62.0±10.5)years(range:22-88 years).According to the location of tumor and the mode of invasion,the tumors were divided into peritoneal type, hepatic type, hepatic hilum type and mixed type, the surgical procedures were divided into regional radical resection and extended radical resection.The correlation between different types and T stage, N stage, vascular invasion, neural invasion, median survival time and surgical procedures were analyzed.Rates were compared by χ(2) test, survival analysis was carried by Kaplan-Meier and Log-rank test. Results: Regional radical resection was performed in 940 cases,including 81 cases in T1 stage,859 cases in T2-T4 stage,119 cases underwent extended radical resection;R0 resection was achieved in 990 cases(93.5%).The overall median survival time was 28 months.There were 81 patients in Tis-T1 stage and 978 patients in T2-T4 stage.The classification of gallbladder cancer in patients with T2-T4 stage: 345 cases(35.3%)of peritoneal type, 331 cases(33.8%) of hepatic type, 122 cases(12.5%) of hepatic hilum type and 180 cases(18.4%) of mixed type.T stage(χ(2)=288.60,P<0.01),N stage(χ(2)=68.10, P<0.01), vascular invasion(χ(2)=128.70, P<0.01)and neural invasion(χ(2)=54.30, P<0.01)were significantly correlated with the classification.The median survival time of peritoneal type,hepatic type,hepatic hilum type and mixed type was 48 months,21 months,16 months and 11 months,respectively(χ(2)=80.60,P<0.01).There was no significant difference in median survival time between regional radical resection and extended radical resection in the peritoneal type,hepatic type,hepatic hilum type and mixed type(all P>0.05). Conclusion: With application of new clinical classification, different types of gallbladder cancer are proved to be correlated with TNM stage, malignant biological behavior and prognosis, which will facilitate us in preoperative evaluation,surgical planning and prognosis evaluation.

TRIM66 promotes malignant progression of hepatocellular carcinoma by inhibiting E-cadherin expression through the EMT pathway.

OBJECTIVE: The aim of this study was to explore the regulatory role of TRIM66 in the development of hepatocellular carcinoma (HCC), and to investigate its underlying mechanism. PATIENTS AND METHODS: A total of 88 pairs of HCC tissues and para-cancerous tissues were surgically resected. The expression of TRIM66 was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The correlation between TRIM66 expression and clinic-pathologic characteristics of HCC patients was analyzed. Follow-up data of enrolled HCC patients were collected for survival analysis. Subsequently, TRIM66 expression in HCC cells was determined by qRT-PCR as well. By constructing si-TRIM66, the biological performances of transfected HCC cells were determined using cell counting kit-8 (CCK-8), colony formation and transwell assay. Western blot was performed to measure the protein expressions of relative genes in epithelial-mesenchymal transition (EMT) pathway. Finally, HCC cells were co-transfected with si-TRIM66 and pcDNA-E-cadherin, followed by detection of invasive and migratory abilities. RESULTS: TRIM66 was highly expressed in HCC tissues compared with that of para-cancerous tissues. High expression of TRIM66 was positively correlated with tumor stage, lymph node metastasis and distant metastasis, whereas not correlated with age and sex of HCC patients. Kaplan-Meier curves revealed that a higher expression of TRIM66 was associated with worse prognosis of HCC. Similarly, TRIM66 was also highly expressed in HCC cells. The knockdown of TRIM66 in HCC cells significantly inhibited the proliferative, invasive and migratory abilities of transfected cells. However, TRIM66 down-regulation significantly induced cell apoptosis. Western blot results showed that TRIM66 knockdown in HCC cells markedly downregulated the protein expressions of E-cadherin, N-cadherin, Vimentin and ß-catenin. The inhibited migration and invasion of HCC cells resulted from TRIM66 knockdown were partially reversed by E-cadherin overexpression. CONCLUSIONS: TRIM66 is highly expressed in HCC, which is positively correlated with tumor stage, lymph node metastasis and distant metastasis of HCC patients. In addition, TRIM66 promotes the malignant progression of HCC by inhibiting E-cadherin through the EMT pathway.

[Spectrum analysis of pathological classification in 463 cases with nasal and paranasal sinuses malignant tumors].

Objective:The characteristics of pathological histological classification of nasal and paranasal sinuses malignant tumors in the past 10 years were analyzed, so as to provide possible basis, direction and ideas for the development of relevant effective treatment measures for nasal and paranasal sinuses malignant tumors in clinical practice. Method:The clinical data of patients with nasal and paranasal sinuses malignant tumors admitted to PLA general hospital from January 2009 to December 2018 were collected. Pathological types were retrospectively analyzed, and disease spectrum distribution, composition ratio and variation tendency of these patients were calculated. Result:Among the 463 patients, the overall pathological types in the top 5 were as follows: squamous cell carcinoma, adenoid cystadenocarcinoma, olfactory neuroblastoma, melanoma, adenocarcinoma. As for male patients, the pathological types in the top 5 were squamous cell carcinoma, adenoid cystic carcinoma, olfactory neuroblastoma, adenocarcinoma, neuroendocrine carcinoma and rhabdomyosarcoma were tied for fifth; the top 5 most common pathological types in female patients were squamous cell carcinoma, adenoid cystic carcinoma, melanoma, rhabdomyosarcoma, and adenocarcinoma. From 2009 to 2013, there were 183 patients with nasal and paranasal sinuses malignant tumors, the top 5 pathological types were squamous cell carcinoma, adenoid cystadenocarcinoma, olfactory neuroblastoma, melanoma, neuroendocrine carcinoma and rhabdomyosarcoma were tied for fifth; From 2014 to 2018, 280 patients with nasal and paranasal sinuses malignant tumors were diagnosed, the top 5 pathological types were squamous cell carcinoma, adenoid cystadenocarcinoma, melanoma, adenocarcinoma, and rhabdomyosarcoma. The ratio of the number of patients from 2009 to 2013 and 2014 to 2018 was about 0.65∶1. Malignant tumors of the nasal and paranasal sinuses tend to occur between the ages of 41 and 60, and the pathological types in the top 5 were squamous cell carcinoma,adenoid cystic carcinoma, adenocarcinoma, melanoma, neuroendocrine carcinoma. Conclusion:Malignant tumors of nasal cavity and sinus were more common in male, and the pathological types such as squamous cell carcinoma, adenoid cystic carcinoma, olfactory neuroblastoma were more common. All age groups have the disease, but the age group of 41-60 years old is the high-risk group of nasal and nasal sinus malignant tumors. However, the incidence rate of melanoma has gradually increased in the past five years, which needs to be paid more attention to.

Recurrent fungal infections in a Chinese patient with CARD9 deficiency and a review of 48 cases.

Deficiency of CARD9 (caspase recruitment domain-containing protein 9) has been reported in individuals with recurrent and invasive fungal infections. We report on a patient who first had Trichosporon asahii affecting the skin then Candida albicans infections involving the digestive tract and knee joint, along with elevated serum IgE. After stimulation with C. albicans, peripheral blood mononuclear cells of this patient produced less tumour necrosis factor-α, interferon-γ and interleukin-17 than those of healthy controls. Furthermore, the serum IgE levels of this patient were positively correlated with the severity of fungal infection during the course of treatment. Sanger sequencing identified one homozygous frameshift mutation (p.D274fsX60) in CARD9. We further performed a review including 48 cases with CARD9 deficiency. According to the data published previously, CARD9-deficient patients demonstrated obviously elevated IgE in serum (median 1300 IU mL-1 ), which could distinguish them from otherwise healthy people with fungal infections (area under the curve 0·94, P < 0·001). Patients carrying the mutations Q289X and Q295X had a higher mortality rate (24% vs. 0%, P < 0·05). Patients with the mutations R18W, R35Q, R70W, G72S or Y91H in the CARD domain, and the nonsense mutation Q295X in the coiled-coil domain, seemed to be more prone to Candida infections (90% vs. 20%, P < 0·005) and central nervous system infections (60% vs. 12%, P < 0·005).

[Clinical and prognostic significance of ABO promotor methylation level in adult leukemia and myelodydysplastic syndrome].

Objective: To investigate the clinical and prognostic significance of ABO promotor methylation level in adult patients with leukemia and myelodydysplastic syndrome(MDS). Methods: ABO promoter methylation level of 182 malignant hematological disease patients and 68 normal controls were detected by bisulfite sequencing PCR.Then clinical features and outcome were compared between hypermethylation group and hypomethylation group. Results: The median methylation rate of ABO promoter in newly diagnosed acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL) were 46.98% and 11.01% respectively, which were both higher than that in controls (2.30%, P<0.05). The methylation rates in remission AML and ALL were 1.58% and 2.30% respectively, which were comparable with that in normal group (P>0.05). As to relapse AML and ALL, methylation rates were 41.26% and 17.50% respectively, also significantly higher than that in controls (P<0.05).In patients with chronic myeloid leukemia (CML) chronic phase, the median methylation rate was 1.00%, which was similar to normal group. But a CML patient who transformed to ALL hadextremely high methylation rate 92.56%. The median methylation rate in patients with MDS significantly elevated as 5.81% compared with that in controls (P<0.05). The median overall survival (OS) of ALL and AML (non-M3) patients with hypermethylation were 12.5 months and 15.3 months, which were significantly shorter than those with hypomethylation (24.0 months and 20.0 months)(P<0.05).The median disease-free survival (DFS) of ALL and AML (non-M3) patients with hypermethylation were 9.9 months and 12.0 months, which were significantly shorter than those with hypomethylation (22.3 months and 18.5 months), (P<0.05). Multivariable analysis suggested that ABO promoter methylation level was an independent predictive factor of OS and DFS in ALL and AML (non-M(3)) patients. Conclusion: ABO promoter hypermethylation is closely related to genesis, development and prognosis of leukemia and MDS. Hypermethylationis related to a clinical poor prognosis compare with hypomethylation.

A novel frameshift mutation of Chediak-Higashi syndrome and treatment in the accelerated phase.

Chediak-Higashi syndrome (CHS) is a rare autosomal recessive immunodeficiency disease characterized by frequent infections, hypopigmentation, progressive neurologic deterioration and hemophagocytic lymphohistiocytosis (HLH), known as the accelerated phase. There is little experience in the accelerated phase of CHS treatment worldwide. Here, we present a case of a 9-month-old boy with continuous high fever, hypopigmentation of the skin, enlarged lymph nodes, hepatosplenomegaly and lung infection. He was diagnosed with CHS by gene sequencing, and had entered the accelerated phase. After 8 weeks of therapy, the boy had remission and was prepared for allogenic stem cell transplantation.

Down-regulation of miR-655-3p predicts worse clinical outcome in patients suffering from hepatocellular carcinoma.

OBJECTIVE: MiR-655-3p has been reported to play important roles in tumor initiation, development, and metastasis in several cancers. This study aimed to assess the potential role of miR-655-3p in the pathogenesis of hepatocellular carcinoma (HCC). PATIENTS AND METHODS: The expression levels of miR-655-3p in HCC tissues were detected by qPCR. The relationship between clinicopathologic characteristics and miR-655-3p was analyzed by chi-square test. Kaplan-Meier curves and multivariate Cox proportional models were used to study the impact on clinical outcome. RESULTS: miR-655-3p was significantly down- regulated in HCC tissues compared to normal adjacent liver tissues (p < 0.01). Low miR-655-3p expression was observed to be closely correlated with positive microvascular invasion, advanced tumor stage and lymph node metastasis (p < 0.05, respectively). The results of Kaplan-Meier analysis showed that patients with high miR-655-3p expression lived shorter than those with low miR-655-3p expression (Log-rank test, p = 0.0002). Multivariate analysis revealed that miR-655-3p was an independent risk factor for HCC (HR=1.533, 95% CI: 0.988-3.891; p = 0.002). CONCLUSIONS: Our data showed that low expression of miR-655-3p was associated with significant characteristics of patients with HCC, and it could function as a potential unfavorable prognostic biomarker.

miR-494 inhibits invasion and proliferation of gastric cancer by targeting IGF-1R.

OBJECTIVE: The aim of this study was to investigate the target gene of miR-494 and its roles in tumor growth of gastric cancer (GC). PATIENTS AND METHODS: Expression of miR-494 was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR) in gastric cancer tissues and cell lines. Then, luciferase reporter assay was used to elucidate whether insulin-like growth factor 1 receptor (IGF1R) is a target gene of miR-494. Finally, the roles and mechanism of miR-494 in the regulation of tumor invasion were further investigated. RESULTS: Relative miR-494 level was found to be significantly lower in patients with GC than healthy controls (p < 0.01). Over-expression of miR-494 could inhibit gastric cancer cell proliferation, migration, and invasion in vitro. Furthermore, we demonstrated that miR-494 binds to the 3'-untranslated region (UTR) of IGF1R and inhibits the expression of the IGF1R protein. CONCLUSIONS: Our data showed that miR-494 acted as a tumor suppressor in GC.

Factors affecting the recurrence and survival of hepatocellular carcinoma after hepatectomy: a retrospective study of 601 Chinese patients.

OBJECTIVE: Indications for resection of hepatocellular carcinoma (HCC) remain controversial. This study aimed to identify prognostic factors that affect overall survival (OS) and disease-free survival (DFS) in patients with HCC after hepatectomy. METHODS: From 2004 to 2010, 601 patients with HCC who underwent resection were enrolled. Factors stratified into the host, biochemical, surgical treatment and tumor-related features in terms of recurrence and overall survival were analyzed. Prognostic factors were evaluated by univariate and multivariate analyses, with Kaplan-Meier survival analyses and Cox proportional hazard model. RESULTS: The overall survival rates of 1-, 3- and 5- year were 79, 62, and 54 %, and the corresponding DFS rates were 51, 38 and 31 %, respectively. In a multivariate analysis, Child-Pugh, serum AFP level, ALT level, time for hepatic resection, tumor differentiation, maximum size of tumors, local necrosis, portal vein tumor thrombus, and TNM Stage were correlated significantly with patients' OS. Gender (P = 0.046), cigarette smoking (P = 0.007), serum AFP level (P = 0.001), GGT level (P = 0.002), maximum size of tumors (P = 0.009), liver cirrhosis (P = 0.025), portal vein tumor thrombus (P = 0.022), microvascular tumor thrombus (P = 0.007) and TNM Stage (P = 0.001) were significantly affected DFS. CONCLUSION: Preoperative AFP level, maximum size of tumors, portal vein tumor thrombus and TNM Stage were revealed as important prognostic factors for OS and DFS through follow-up of a relatively large cohort of Chinese HCC patients.

Impaired function of CD19(+) CD24(hi) CD38(hi) regulatory B cells in patients with pemphigus.

BACKGROUND: Pemphigus is an organ-specific autoimmune bullous disease. OBJECTIVES: To determine the role of regulatory B cells (Bregs) in patients with pemphigus. METHODS: The frequency of the occurrence of CD19(+) CD24(hi) CD38(hi) Bregs was detected from 34 patients with pemphigus and 20 healthy controls. Interleukin (IL)-10 secretion was processed after stimulating B cells. Specific antidesmoglein antibody (Ab) titres and their subclasses were also measured. Ab response and cytokine production from peripheral blood mononuclear cells (PBMCs) with or without Bregs were analysed. RESULTS: The number of Bregs was significantly increased in patients with pemphigus compared with healthy controls (15 ± 7% vs. 9 ± 3%; P < 0·01) and the proportion of Bregs in the active groups (newly diagnosed and chronic active patients) was significantly higher than in remittent individuals (16 ± 7% vs. 13 ± 8%; P = 0·04). The IL-10-producing B cells were significantly increased upon stimulation both in patients and in healthy controls. However, the increase ratio of IL-10-producing B cells between short- and long-term stimulation was significantly lower in patients with pemphigus (1·0-fold vs. 2·6-fold increase in control group; P < 0·01). Strikingly, Bregs from the controls were able to suppress interferon (IFN)-γ expression and T helper cell 1 (Th1) immune response (26% inhibition rate), while the suppressive function of Bregs from patients with pemphigus was significantly decreased (9% inhibition rate). There was no difference in Ab levels from PBMCs with or without Bregs after stimulation. CONCLUSIONS: Bregs in patients with pemphigus are elevated but with defective regulatory function on Th1 cells.

Effects of prolonged intensive lipid-lowering therapy on the characteristics of carotid atherosclerotic plaques in vivo by MRI: a case-control study.

High-resolution magnetic resonance imaging (MRI) with flow suppression not only provides useful information on luminal and wall areas of the carotid artery but also can identify the principal tissue components of the carotid atherosclerotic plaque. The effects of intensive lipid-lowering therapy on these MRI tissue characteristics were examined in patients with coronary disease (CAD). Eight CAD patients who have been receiving intensive lipid-lowering treatment (niacin 2.5 g/d, lovastatin 40 mg/d, and colestipol 20 g/d) for 10 years in the Familial Atherosclerosis Treatment Study (FATS) follow-up were randomly selected from among 60 such treated patients. Eight CAD patients who were matched to the treated patients for age (+/-3 years), baseline low density lipoprotein (+/-5 mg/dL), and triglycerides (+/-50 mg/dL) but who had never been treated with lipid-lowering drugs were selected as controls. For each of these 32 carotid arteries, luminal and plaque areas were measured by planimetry, in a blinded protocol, from the magnetic resonance image that showed most plaque. Fibrous tissue, calcium, and lipid deposits were identified on the basis of established criteria. Plaque composition was estimated as a fraction of total planimetered area. Patients treated with 10-year intensive lipid-lowering therapy, compared with control subjects, had significantly lower low density lipoprotein cholesterol levels (84 versus 158 mg/dL, respectively; P<0.001) and higher high density lipoprotein cholesterol levels (51 versus 37 mg/dL, respectively; P<0.001). As a group, treated patients, compared with untreated control subjects, had a smaller core lipid area (0.7 versus 10.2 mm(2), respectively; P=0.01) and lipid composition (1% versus 17%, respectively). Group differences in luminal area (55 [treated] versus 44 [control] mm(2), P=NS) and plaque area (58 [treated] versus 64 [control] mm(2), P=NS) tended to favor treatment. MRI appears useful for estimating carotid plaque size and composition. Hyperlipidemic CAD patients frequently (97%) have at least moderate (>/=40% area stenosis) carotid plaque. In this case-control study, prolonged intensive lipid-lowering therapy is associated with a markedly decreased lipid content, a characteristic of clinically stable plaques.

Antioxidant supplements block the response of HDL to simvastatin-niacin therapy in patients with coronary artery disease and low HDL.

One strategy for treating coronary artery disease (CAD) patients with low HDL cholesterol (HDL-C) is to maximally increase the HDL-C to LDL-C ratio by combining lifestyle changes with niacin (N) plus a statin. Because HDL can prevent LDL oxidation, the low-HDL state also may benefit clinically from supplemental antioxidants. Lipoprotein changes over 12 months were studied in 153 CAD subjects with low HDL-C randomized to take simvastatin and niacin (S-N), antioxidants (vitamins E and C, beta-carotene, and selenium), S-N plus antioxidants (S-N+A), or placebo. Mean baseline plasma cholesterol, triglyceride, LDL-C, and HDL-C levels of the 153 subjects were 196, 207, 127, and 32 mg/dL, respectively. Without S-N, lipid changes were minor. The S-N and S-N+A groups had comparably significant reductions (P

Electron-beam tomography coronary calcium scores are superior to Framingham risk variables for predicting the measured proximal stenosis burden.

Previous studies of electron-beam tomography (EBT) have correlated coronary calcium scores with simplistic visual estimates of disease severity. In a clinical trial designed to evaluate 2 treatment strategies in coronary artery disease (CAD) patients with low levels of high-density lipoprotein cholesterol, we used quantitative coronary angiography to measure composite proximal stenosis burden from the baseline coronary angiogram and assessed the traditional Framingham risk variables in 146 patients. Stenosis burden is the sum, per patient, of percent stenosis for the worst lesion found in each of 9 standard proximal coronary segments. EBT estimates of coronary calcium (Agatston score, calcium volume score) were obtained for 115 of these patients. Stenosis burden was correlated with the calcium scores and risk variables. The best traditional correlates of stenosis burden were smoking status (r = 0.31, p = 0.001), prior myocardial infarction (r = 0.24, p = 0.005), body mass index (r = 0.23, p = 0.005), pack-years smoking (r = 0.20, p = 0.05), and age (r = 0.17, p = 0.04). With adjustment for age, all these correlations improved (eg, body mass index x age [r = 0.28, p = 0.001]). In addition, total cholesterol x age (r = 0.22, p = 0.008), fibrinogen x age (r = 0.19, p = 0.03), and systolic blood pressure x age (r = 0.18, p = 0.03) became significant correlates. Spearman correlations of the calcium scores with stenosis burden were considerably greater (Agatston: r = 0.62, p <0.0001; calcium volume: r = 0.63, p <0.0001). In multivariate regression analysis, calcium score, body mass index, and history of myocardial infarction were independent correlates of stenosis burden (R(2) = 0.45). At a given point in time, the EBT coronary calcium scores are greatly superior to the Framingham risk factors in predicting the measured proximal stenosis burden. Agatston and calcium volume scores are comparably predictive of stenosis burden.