Objective To establish a real-time PCR based method for detection of survivin gene samples methylation with dye SYBR Green Ⅰ.Methods DNA samples from 25 pairs of gastric cancer tissue and matched normal gastric tissues were digested with mCpG-sensitive Hpa Ⅱand Msp Ⅰ, and detected by SYBR Green Ⅰ real-time PCR for survivin exon 1 specific primers. The concentration of digested DNA was monitored by real-time quantitative PCR with survivin intron 2 specific primers. A normal genomic DNA which served as a standard was diluted serially 10-fold to determine the sensitivity of real-time PCR. The products were further verified by agarose gel electrophoresis analysis.Results After HpaⅡ enzyme digestion, a peak of PCR product derived from methylated targeted gene was found in the melting curve at Tm (91.5±0.5) ℃, which was verified as 338 bp strap. The control gene (survivin intron 2) characteristic of Tm (79.5±0.5) ℃ was amplified in all digested samples, indicating no non-specific degradation occurred in the genome DNA. The sensitivity of this method was 100 copies/μL. The new established PCR system was applied to detecting 25 cases of lung cancer tissue samples. As a result, the frequency of survivin exon 1 demethylation was 96%.Conclusion As a reliable new method for detection of survivin exon 1 methylation, SYBR Green Ⅰ real-time fluorescent PCR assay is rapid, accurate, sensitive, timing and simple.

Survivin variants specific real time quantitative RT-PCR was developed to analyze their expression in 53 paired cancer and para-cancerous tissues, and the expression of the wild-type survivin protein was detected by immunohistochemistry. The results showed that survivin mRNA and protein were expressed in gastric cancer and para-cancerous tissues. The survivin-2B was dominantly expressed in para-cancerous tissues, whereas the survivin-DeltaEx3 was more frequently detected in cancer tissues. The positive rate of survivin-2a was 100% in both cancer and para-cancerous tissues, but its relative transcript expression level was not significantly increased in cancer tissues in comparison with para-cancerous tissues. The correlation analysis revealed that the expression of survivin-2a mRNA was significantly associated with that of total survivin (r (s)=0.4178, P=0.0018), whereas inversely to that of survivin-DeltaEX3 (r (s)=-0.4506, P=0.0007). It was suggested that survivin-2a may act as an antagonist of survivin-DeltaEX3. The balance between antiapoptotic survivin iso-forms and nonantiapoptotic ones may play an important role in tumorigenesis and tumor progression. Promising value is hinted to analyze survivin and its variants in tumor early diagnosis and distinguishing malignant tumors from benign ones.
Microtubule-Associated Proteins/genetics ; Microtubule-Associated Proteins/*metabolism ; Protein Isoforms/genetics ; Protein Isoforms/metabolism ; RNA, Messenger/genetics ; RNA, Messenger/metabolism ; Stomach Neoplasms/*metabolism ; Stomach Neoplasms/pathology ; Tumor Cells, Cultured ; Tumor Markers, Biological/genetics ; Tumor Markers, Biological/*metabolism

Objective: To discuss the way of early nutritional support in the patients after the liver transplantation. Methods: The nutritional support was used in 28 patients who had suffered the orthopetic liver transplantation (OLT).Parenteral nutrition was started as soon as possible after the operation. On the fourth day after the operation, enteral nutrition was also used. Results: 5 patients died from the postoperative complications. 23 patients survived and had been in better nutrition. Conclusion: The early nutrition support can play an important role in the patients recovery after OLT. EN should be administrated immediately when the gut functions.

Objective To investigate the correlations of expression of interferon inducible protein 10 ( IP-10) mRNA in peripheral blood mononuclear cells ( PBMCs) with serum levels of HBsAg and HBV DNA in patients with HBV-related acute on chronic liver failure ( HBV-ACLF ) , and to assess its value in predicting disease prognosis .Methods Eighty patients with HBV-ACLF, 60 patients with chronic hepatitis B (CHB), and 25 healthy subjects were enrolled during October 2013 and February 2015.IP-10 mRNA in PBMCs was measured by real time quantitative polymerase chain reaction ( RT-qPCR) , and the difference in IP-10 mRNA expression among three groups was compared by ANOVA .Serum levels of HBsAg and HBV DNA were also measured in patients with HBV-ACLF, and Pearson correlation test was performed to analyze the correlations of IP-10 mRNA with HBsAg and HBV DNA levels .t test was used to analyze the differences in IP-10 mRNA, HBsAg and HBV DNA levels between fatal and surviving cases after 3-month entecavir therapy in HBV-ACLF group.Receiver operating characteristic curves ( ROC) was used to evaluate the prognostic value of IP-10 mRNA, HBsAg and HBV DNA in HBV-ACLF patients .Results IP-10 mRNA level in HBV-ACLF group was 0.998 ±0.186, which was higher than those in CHB patients and healthy controls (0.641 ±0.083 and 0.412 ±0.062, t=3.841 and 16.661, P<0.01).Pearson correlation analysis showed that IP-10 mRNA level was negatively correlated with HBsAg level in HBV-ACLF patients (r=-0.576, P<0.01), but positively correlated with HBV DNA level (r=0.547, P<0.01).After 3-month entecavir therapy , IP-10 mRNA level in surviving cases of HBV-ACLF group was 0.894 ±0.164, which was lower than that in fatal cases ( 1.103 ±0.177, t =-4.328, P <0.01 ); HBsAg level in surviving cases was higher than that in fatal cases (3.303 ±0.565 vs.2.605 ±0.844, t =3.251, P<0.01).The area under ROC of IP-10 mRNA in evaluating prognosis of HBV-ACLF was 0.820, which was higher than those of HBsAg (0.663) and HBV DNA (0.570).Conclusions IP-10 mRNA in patients with HBV-ACLF is over-expressed and is correlated with HBsAg and HBV DNA levels .It may be used for predicting the prognosis of patients with HBV-ACLF.

0.05) between two groups. Abdominal distention and diarrhea occurred in 6 patients in observed group. Nitrogen balance, PA, TFN, IgA, IgG and IgM after nutritional support were better than those before nutritional support in two groups. Nitrogen balance, PA, TFN, IgA, IgG and IgM were better in observed group than those in the control group. Conclusions:Early nutritional support can improve metabolic status,the immune function and the nitrogen balance in brain operation patients.

Objective To investigate the expression of interferon inducible protein-10 (IP-10) in peripheral blood mononuclear cells (PBMC) of patients with hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF), and its correlation with disease severity.Methods Eighty patients with HBV-ACLF, 60 patients with chronic hepatitis B (CHB), and 25 healthy controls were enrolled from the Affiliated Hospital of Hubei University of Arts and Science during October 2013 and February 2015.IP-10 mRNA in PBMC was measured by real time quantitative PCR.Independent sample t test was used to analyze the difference in IP-10 mRNA expression between HBV-ACLF patients with model for end-stage liver disease (MELD) ＜ 30 and ≥30, and Pearson correlation test was performed to analyze the correlations of IP-10 mRNA expression with alanine aminotransferase (ALT), total bilirubin (TBil) and international normalized ratio (INR).Results The expressions of IP-10 mRNA in HBV-ACLF patients was 1.00 ± 0.19, which was higher than those in CHB patients and healthy controls (0.64 ± 0.08 and 0.41 ± 0.06, t =3.841 and 16.661, all P ＜ 0.01).The expression of IP-10 mRNA in HBV-ACLF patients with MELD ＜ 30 was 0.96 ±0.19, which was lower than that in patients with MELD ≥ 30 (1.14 ± 0.21, t =-2.283, P ＜0.05).Pearson correlation analysis showed that IP-10 mRNA level in HBV-ACLF patients was positively correlated with ALT, TBil and INR (r =0.697, 0.738 and 0.775, all P ＜ 0.01).Conclusion IP-10 mRNA is over-expressed in PBMC of patients with HBV-ACLF, and it is correlated with disease severity, which suggests that IP-10 may play an important role in the progression of liver failure.

Objective To investigate the predictive value of dynamic changes of interferon-inducible protein-10 (IP-10) expression in the peripheral blood mononuclear cells and the model for end-stage 1iver disease (MELD) scores for short-term mortality in hepatitis B virus related acute-on-chronic liver failure (HBV-ACLF) patients .Methods Eighty patients with HBV-ACLF admitted to the Affiliated Hospital of Hubei College of Arts and Sciences from October 2013 to August 2015 were selected .During 3 months of follow-up ,33 patients died and 47 survived .The expression level of IP-10 and MELD score of two groups were measured at admission and week 1 and week 2 after treatment .The means between two groups were compared .Accuracy of predicting short-term mortality was performed by area under receiver operating characteristic curve (AUC) .Multivariate logistic regression analysis and Kaplan-Meier survival curve were used to analyze the effect of IP-10 expression and MELD score on the mortality of HBV-ACLF patients . Results The expressions of IP-10 at admission and at week 1 and 2 after treatment in the death group were 1 .095 ± 0 .202 ,1 .071 ± 0 .181 ,and 1 .078 ± 0 .198 ,respectively ,those in the survival group were 0 .894 ± 0 .181 ,0 .770 ± 0 .153 ,and 0 .732 ± 0 .137 ,respectively ,which were significantly different (t =4 .66 ,8 .02 and 9 .27 ,respectively ,all P < 0 .01) .The MELD scores at admission and at week 1 and 2 after treatment in the death group were 26 .70 ± 5 .50 ,27 .39 ± 6 .24 ,and 28 .64 ± 6 .44 ,respectively , those in the survival group were 23 .89 ± 4 .41 ,21 .57 ± 4 .68 ,and 18 .87 ± 3 .92 ,respectively ,which were significantly different (t= 2 .53 ,4 .77 and 8 .42 ,respectively ,all P< 0 .01) .Analysis of variance showed that the MELD score and IP-10 expression in the survival group at admission were significantly higher than those at week 1 and week 2 after treatment (F= 13 .464 and 15 .711 ,respectively ,both P< 0 .01) ,while there were no significant differences in the death group (F = 0 .129 and 0 .864 ,respectively ,both P >0 .05) .The AUC of IP-10 at week 2 after treatment was 0 .935 ,that of MELD score was 0 .903 (Z =0 .788 ,P= 0 .045) ,while there was no significant difference of AUC between week 1 and week 2 (0 .935 vs 0 .909 ,Z = 0 .640 ,P> 0 .05) .In addition ,the AUC of IP-10 level at week 1 and MELD score at week 2 after treatment showed no significant difference (0 .909 vs 0 .903 ,Z = 0 .133 , P > 0 .05) .Logistic multivariate regression analysis showed that IP-10 ≥ 0 .902 at week 1 ,MELD ≥ 22 .5 and IP-10 ≥ 0 .846 at week 2 were independent risk factors for death (OR= 11 .29 ,6 .60 ,and 15 .27 ,respectively ;95％ CI =1 .06 - 119 .74 ,1 .27 - 34 .26 ,and 1 .39 - 167 .62 ,respectively ;all P< 0 .05) .Conclusion The dynamic monitor of both IP-10 levels and MELD scores may have greater value in predicting prognosis of patients with HBV-ACLF .

Survivin variants specific real time quantitative RT-PCR was developed to analyze their expression in 53 paired cancer and para-cancerous tissues, and the expression of the wild-type survivin protein was detected by immunohistochemistry. The results showed that survivin mRNA and protein were expressed in gastric cancer and para-cancerous tissues. The survivin-2B was dominantly expressed in para-cancerous tissues, whereas the survivin-△Ex3 was more frequently detected in cancer tissues. The positive rate of survivin-2a was 100% in both cancer and para-cancerous tissues,but its relative transcript expression level was not significantly increased in cancer tissues in comparison with para-cancerous tissues. The correlation analysis revealed that the expression of survivin-2a mRNA was significantly associated with that of total survivin (rs=0.4178, P=0.0018),whereas inversely to that of survivin-/EX3 (rs=-0.4506, P=0.0007). It was suggested that survivin-2a may act as an antagonist of survivin-△EX3. The balance between antiapoptotic survivin iso-forms and nonantiapoptotic ones may play an important role in tumorigenesis and tumor progression. Promising value is hinted to analyze survivin and its variants in tumor early diagnosis and distinguishing malignant tumors from benign ones.

OBJECTIVETo investigate the clinical effect of repairing soft tissue defect after hand wound using reverse island skin flap of upper carpal cutaneous branches of ulnar artery.

METHODSFrom June 2010 to November 2016, 12 patients with hand soft tissue defects were repaired by reverse island skin flap of upper carpal cutaneous branches of ulnar artery, including 9 males and 3 females with an average age of (35.2±9.4) years old ranging from 22 to 58 years. The defect area varied from 7.0 cm×3.0 cm to 12.0 cm×7.0 cm. Time interval from injury to operation ranged from 3 to 15 days with an average of (8.4±2.6) days. The flap was designed beforehand according to the size of the defect, sharply dissected the aponeurotic fascia from the proximal to the distal, abscised the communicating branch between the flap and the ulnar artery at the wrist epithelial branch, repairing the defect of flap with method of metastasis retrograde. The sensation, shape of the flap and hand function were observed, and the upper extremity function was evaluated according to the standard of hand surgery branch from Chinese Medical Association to assessment of functional recovery.

RESULTSThe flaps in 10 patients obtained primary healing, the healed time was 14 to 18 days with an average of(15.0±1.5) days. Two patients occurred distal flap necrosis, and wound surface healed after change dressing and skingrafting cover. All patients were followed up from 3 to 15 months with an average of(7.0±3.8) months. According to the upper extremity functional evaluation standard by hand surgery branch of Chinese Medical Association, 2 cases got excellent results, 7 good, 2 fair and 1 poor.

CONCLUSIONSReverse island skin flap of upper carpal cutaneous branches of ulnar artery for the treatment of soft tissue defect caused by hand wound has advantages of concealed donor area, no need sacrifice the main blood vessel, flap thin and no need repair it for thick and thin.