OBJECTIVETo screen long non-coding RNA which influences radiosensitivity of colorectal carcinoma cell lines and investigate the mechanism.

METHODSUnder different doses of radiation, colony formation assay and single-hit multi-target model were conducted to draw dose-survival curve and SF2 value of colorectal carcinoma cell lines(RKO, Lovo) was calculated. High-throughput lncRNA/mRNA chips were used to screen lncRNA genes and protein coding genes with expression differences more than 2 folds between RKO, Lovo cell lines and RKO cell line receiving 2Gy radiation. The main action pathway was computed by Gene Ontology analysis combined with Pathway analysis in order to explore the mechanism which induces the effect of lncRNA on radiosensitivity of colorectal carcinoma cell lines. Further experiment on P53, P21, cyclin D1 expression contents of RKO cell line was confirmed by real-time RT-PCR.

RESULTSLovo(SF2=0.47) was more sensitivity to radiation than RKO(SF2=0.53) according to the outcome of colony formation assay. High-throughput lncRNA/mRNA chips identified a total of 268 lncRNA genes and 270 protein coding genes. Gene Ontology analysis showed that the expression of genes associated with cell cycle process were significantly different (38.6%). There was a significant relationship between expression of several lncRNAs and CCND1 gene. Real-time RT-PCR showed no significant differences of P53 and P21 expression in RKO and Lovo cell lines(P>0.05), while cyclin D1 expression of RKO cell line was higher than that of Lovo cell lines(P<0.05). After exposed to 2 Gy doses of radiation, there was an obvious decrease of cyclin D1 expression in RKO cell lines(P<0.05), while P53 and P21 expressions were not different(P>0.05).

CONCLUSIONThe possible mechanism is that lncRNAs compose transcription compound to combine with CCND1 gene and influence radiosensitivity of colorectal carcinoma cell lines by regulating expression of cyclin D1, which is independent of P53-P21-cyclin D1 pathway.

Cell Line, Tumor ; Colorectal Neoplasms ; metabolism ; pathology ; Cyclin D1 ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; RNA, Long Noncoding ; Radiation Tolerance

OBJECTIVETo investigate the effect of multidrug resistance-associated protein 4 (MRP4) expression on the radiosensitivity of colorectal carcinoma cell lines in vitro.

METHODSThe vector of shRNA for RNA interference was constructed and then transfected into HCT116 cell line to steadily down-regulate the expression of MRP4. HCT116 cells were divided into 3 groups including the CON group(non-transfected), NC group (negative control virus was added), and KD group (RNAi target was added for transfection). To test the effectiveness of RNA interference, real-time polymerase chain reaction and Western blot were used to measure the expression pattern of MRP4 at both mRNA and protein levels, respectively. For the examination of the effect of RNA interference of MRP4 on the radiosensitivity, flow cytometry was used to calculate the rate of apoptotic cells 24 h after 4 Gy radiation. Proliferation of the cells was measured via MTT assay at different time points.

RESULTSShRNA plasmid was successfully constructed. Transfection of this constructed vector into HCT116 cell line caused steady silencing of MRP4 expression (HCT116-KD). MRP4 mRNA and protein expression were significantly down-regulated following RNA interference(P<0.05). Twenty-four hours after radiation, the apoptosis rate of KD cell line was (71.7±0.8)%, significantly higher than that in the CON group [(56.1±0.9)%] and NC group[(59.8±0.8)%](P<0.05). Fourty-eight hours and 72 hours after radiation, the proliferation was significantly inhibited in KD cells compared to the control groups(P<0.05).

CONCLUSIONSExpression of MRP4 is closely related to radio-tolerance of colorectal carcinoma. Down-regulation of MRP4 expression by RNA interference enhances radiosensitivity of colorectal carcinoma cell lines in vitro. MRP4 may be an effective molecular marker for predicting the radiosensitivity of colorectal carcinoma.

Colorectal Neoplasms ; genetics ; metabolism ; Down-Regulation ; HCT116 Cells ; Humans ; Multidrug Resistance-Associated Proteins ; genetics ; RNA Interference ; Radiation Tolerance ; genetics

OBJECTIVETo investigate the involvement of PI3K/Akt signaling pathway in the changes of urine protein in adriamycin-induced nephropathic rats treated with dexamethasone and LY294002 (a PI3K inhibitor).

METHODSSD rats were randomized into normal control group, ariamycin-induced nephropathy group (ADR group), ariamycin+dexamethasone group (DEX group), and ADR+DEX+LY294002 group (LY294002 group). On days 7, 14 and 28 after the treatments, 24-h urine was collected from the rats to analyze the total urine proteins. The renal tissues were obtained on day 28 to examine the expressions of p-AKT, AKT and Bad proteins in the cortical tissues using Western blotting; the expression of Bad mRNA in the cortical tissues was measured by QPCR.

RESULTSUrine protein increased progressively in ADR group accompanied by significantly reduced p-AKT/AKT ratio and increased Bad mRNA expression in comparison with those in the normal control group (P<0.05). Urine protein was obviously reduced in DEX group with comparable p-AKT/AKT ratio and Bad mRNA expression level with those in the control group (P>0.05). Urine protein showed no significant reduction in LY294002 group, but the p-AKT/AKT ratio was significantly reduced and Bad mRNA expression was increased compared with those in DEX group (P<0.05).

CONCLUSIONDexamethasone increases the expression of Bad mRNA and reduces urine protein in adriamycin-induced nephropathic rats by activating PI3K/Akt signaling pathway. LY294002 can inhibit PI3K/Akt signaling pathway to block the effect of dexamethasone, suggesting that PI3K/Akt signaling pathway is one of the signaling pathways that mediate the effect of dexamethasone on proteinuria.

Purpose::The reconstruction of Allen's type IV fingertip amputation is a clinical challenge. Our team designed bilateral unequal-sized hallux osteo-onychocutaneous free flaps for the long-term reconstruction of Allen's type IV fingertip amputation and conducted a retrospective study with a 5-year follow-up aims to evaluate the effects of this technique.Methods::A retrospective analysis with a 5-year follow-up including 13 patients with Allen's type IV fingertip amputation who were admitted to our hospital from January 2010 to January 2017 was conducted. The patients were treated with bilateral unequal-sized hallux osteo-onychocutaneous free flaps. The operation time, intraoperative blood loss, and complications were recorded, and the survival rate of the transplanted flaps was calculated. During the 5-year follow-up after operation, the nail growth time was recorded and the finger appearance was observed. At the last follow-up appointment, the length, width, and girth of the reconstructed fingertip and contralateral normal fingertip, range of motion of the reconstructed fingertip and contralateral normal fingertip, Semmes-Weinstein test (for the evaluation of tactile sensation), and two-point discrimination testing results were recorded. SPSS 22.0 software was used for the statistical analysis and the data are presented as mean ± SD.Results::The mean operation time was (5.62 ± 0.51) h, the mean intraoperative blood loss was (34.15 ± 3.13) mL, and the survival rate of the transplanted flaps was 100%. During the 5-year follow-up, the average nail growth time was (10.14 ± 1.98) months and the average bone union time was (3.78 ± 0.91) months. The length, width, and girth of the reconstructed fingertip were (31.52 ± 3.73) mm, (17.82 ± 1.74) mm, and (59.75 ± 3.04) mm, respectively, which did not differ from those of the contralateral normal fingertip. The range of motion of the reconstructed fingertip was (12.15 ± 2.79) degrees which is different from that of the contralateral normal fingertip. The average tactile sensation evaluated via the Semmes-Weinstein test and the average two-point discrimination test of the reconstructed fingertip were (0.39 ± 0.17) g and (7.46 ± 1.14) mm, respectively, which were not different from those of the contralateral normal fingertip. The average Maryland score of feet in the donor area was 87.66 ± 7.39, which was satisfactory.Conclusion::Bilateral unequal-sized hallux osteo-onychocutaneous free flaps are an effective method to reconstruct Allen's type IV fingertip amputations with a satisfactory appearance and good sensory function.

Objective To observe the clinical effect of dynamic blood glucose monitoring combined with intensive insulin therapy in critically ill patients with hyperglycemia in intensive care unit (ICU).Methods A total of 135 cases of critically ill patients with severe hyperglycemia were randomly divided into control group (n =67) and treatment group (n =68).Control group was given fingertip blood glucose monitoring,3 times a day combined with syringe pump continuous intravenous 2-4 U · h-1 insulin.Treatment group was given dynamic blood glucose monitoring (every 6 h on blood glucose changes recorded) combined with insulin intensive treatment (when patients' initial blood glucose ＞ 12.1 mmol · L-1,the patients were intravenous infusion 4 U · h-1 insulin,if the blood glucose did not decline,the appropriate increase in 1.5-2.0 U · h-1 insulin).Two groups were treated continuously for 2 weeks.The fasting blood glucose (FBG),ICU hospitalization time,mean mechanical ventilation time,mortality and hypoglycemia were observed in two groups.Results After 3,5,7 d treatment,the FBG in treatment group were (6.34 ± 1.48),(5.73 ± 1.23),(5.24 ± 0.86) mmol· L-1,had significant difference with those in control group,which were (8.76 ±2.36),(8.46 ±2.19),(7.59 ±2.19) mmol · L-1(P ＜0.05).The ICU hospitalization time,mean mechanical ventilation time in treatment group were (8.63 ±5.72),(6.25 ±4.61) d,had no significant difference with those in control group,which were (9.78 ±6.34) and (6.75 ±4.78) d (P ＞0.05).There were 7 cases (10.29％) of mortality in treatment group and 11 cases (16.41％) in control group,with no significant difference (P ＞ 0.05).There were 6 cases (8.82％) of hypoglycemia in treatment group and 14 cases (20.89％) in control group,with no significant difference (P ＜ 0.05).Conclusion In the intensive care unit,hyperglycemia patients with dynamic blood glucose monitoring combined with insulin intensive treatment can more accurately grasp hyperglycemia critically ill patients' blood glucose changes,and then more scientific development of insulin use,can quickly reduce the patients' blood glucose and reduce the incidence of hypoglycemia,has a very good clinical efficacy.