Simultaneous quantification of two steroidal glycosides after oral gavage of Marsdenia tenacissima extract in rats using a LC-MS/MS method.

A specific, sensitive and accurate analytical LC-MS/MS assay was developed for the simultaneous determination of two steroidal glycosides, tenacissoside H and tenacissoside I, in rat plasma. An Agilent ZORBAX SB-C18 column was used with an isocratic mobile phase system composed of methanol-water-formic acid (70:30:0.1, v/v/v) at a flow rate of 0.3 mL/min. The analysis was performed on a positive ionization electrospray mass spectrometer via selected reaction monitoring mode scan. One-step protein precipitation with acetonitrile was chosen to extract the analytes from plasma. The lower limits of quantification were 0.9 ng/mL for tenacissoside H and tenacissoside I. The intra- and inter-day precisions were 2.03-11.56 and 3.76-11.62%, respectively, and the accuracies were <110.28% at all quality control levels. The validated method was applied to a pharmacokinetic study in rats after oral gavage of Marsdenia tenacissima extract.

miR-125b regulates side population in breast cancer and confers a chemoresistant phenotype.

Resistance to chemotherapy is a major obstacle for the effective treatment of breast cancer and is partially due to the presence of drug resistant stem cell-like side population (SP). Previous studies have shown elevated miR-125b is associated with chemoresistance and metastasis; however, the relationship between miR-125b and SP cells remains unknown. In this study, we isolated and characterized SP cells in a panel of breast cancer cell lines and primary cancer cells from breast cancer patients. SP cells showed cancer stem cells (CSCs) properties, including self-renewal, resistance to chemotherapy and high expression of stem cell markers. The percentage of SP cells was higher in chemotherapy resistant patients compared to that in chemotherapy responsive patients (5.8 ± 2.4% in non-responsive patients vs. 1.2 ± 0.5% in responsive patients, P = 0.012). Importantly, SP cells had higher level of miR-125b than NSP cells and the elevated miR-125b expression in chemoresistant cancer cells were due to high percentage of SP cells. Overexpression of miR-125b correlated with an increase in tumor SP and CSC property, whereas knockdown of miR-125b correlated with decreased incidence of SP. In addition, miR-125b overexpression in breast cancer cells induced epithelial-mesenchymal transition (EMT)-like cellular marker alteration, suggesting a potential mechanism of miR-125b in the regulation of cancer stem-like SP cells. Taken together, these results suggest an important role for miR-125b in breast cancer chemoresistance by maintaining cancer stem-like SP fraction, and raise the possibility that miR-125b may be a significant prognostic response marker for cancer therapy.

Blocking the IGF2BP1-promoted glucose metabolism of colon cancer cells via direct de-stabilizing mRNA of the LDHA enhances anticancer effects.

Colorectal cancer (CRC) is a commonly diagnosed cancer with poor prognosis and high mortality rate. Hyperthermia (HT) is an adjunctive therapy to enhance the antitumor effects of traditional chemo- or radio- therapy. Here, we report that a cluster of essential regulator genes and speed-limit enzymes of glucose metabolism were significantly elevated under HT from a glucose metabolism PCR array analysis. Under low glucose supply or glucose metabolism inhibition, CRC cells displayed increased sensitivity to HT treatments. By transcript sequencing from the established HT resistant (HTR) colon cancer cell line LoVo HTR, we observed that IGF2BP1, an RNA-binding protein, was significantly upregulated in HTR cells compared with parental cells. Furthermore, LDHA mRNA was identified as an IGF2BP1 direct target. An RNA immunoprecipitation assay and RNA pull-down assay consistently illustrated IGF2BP1 specifically bonds to the 3' UTR of LDHA mRNA, leading to enhanced stability of LDHA mRNA. Finally, we demonstrated that inhibiting the IGF2BP1-promoted glycolysis sensitized colon cancer cells to HT treatment via both in vitro and in vivo experiments. Our findings suggest that targeting the IGF2BP1-LDHA-glycolysis pathway might be a promising therapeutic approach to enhance the anti-cancer effects of HT treatment.

[Extracellular signal-regulated protein kinase activation in endometrium with polycystic ovary syndrome and its significance].

OBJECTIVE: to investigate the activation of mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase (ERK) signaling pathway in the endometrium of women with polycystic ovary syndrome (PCOS) and its effect and significance in the cause of hyperplasia and carcinoma; and investigate the factors which affect the activation of the MAPK/ERK signaling pathway. METHODS: collected 52 patients diagnosed as PCOS who were taken dilation and curettage of uterus as study, while 32 non-PCOS patients matched as control group. Serum hormonal parameters, fasting blood glucose and insulin were measured in all patients. The PCOS patients were sub-group as insulin resistance group and non-insulin resistance group; all the patients were carried out pathology inspection of endometria, and the PCOS patients were sub-group as endometrial hyperplasia and carcinoma group and normal endometrium group based on the outcome of pathology inspection. Western blot were performed to detect the expressions of ERK1/2 and phosphorylated ERK1/2 (p-ERK1/2), the activation of ERK1/2. RESULTS: (1) the expression of p-ERK1/2 [(61 ± 13)%] in the endometrium in PCOS group was higher than that in the control [(44 ± 10)%, P < 0.01]. (2) The expression of p-ERK1/2 was significantly increased in group of endometrial hyperplasia and carcinoma [(70 ± 11)%] compared to that in group of normal endometrium [(55 ± 10)%, P < 0.01], while there were significant difference between group of insulin resistance [(63 ± 13)%] and group of non-insulin resistance [(55 ± 7)%, P < 0.01]. (3)Fasting insulin level, insulin area under the curve and body mass index were related to the expression of p-ERK1/2 in endometrium with PCOS, the correlation coefficient were 0.447, 0.456 and 0.381, respectively (all P < 0.01). CONCLUSIONS: the MAPK/ERK signaling pathway in endometrium with PCOS was overactivation, which was related to the endometrial hyperplasia and carcinoma; while the activation of MAPK/ERK signaling pathway were effected by insulin resistance and hyperinsulinemia.

[Comparison of transurethral surgical methods for treating small-size prostate hyperplasia].

OBJECTIVE: To analyze different transurethral surgical methods for the treatment of small-size benign prostate hyperplasia (BPH) in order to improve the curative effect. METHODS: The clinical data of 52 cases of small-size BPH treated by transurethral surgery were reviewed and analyzed. Of the total number, 12 underwent transurethral prostate resection (TURP), 18 TURP plus transurethral incision of the bladder neck (TUIBN) and 22 TURP plus transurethral resection of the bladder neck (TURBN). The curative effect of the three different surgical methods was evaluated by international prostate symptom score (IPSS), maximum flow rate (Qmax) and post-voiding residual urine volume (PVR). RESULTS: In the TURP group, 3 cases were complicated with contracture of the bladder neck, and the IPSS, Qmax and PVR were (12.2 +/- 3.2), (11.7 +/- 2.6) ml/s and (27.6 +/- 13.0) ml, respectively. In the TURP + TUIBN group, there was only 1 case of the complication and the three indices were respectively (8.6 +/- 3.2), (16.7 +/- 3.0) ml/s and (20.0 +/- 8.0) ml. No complication was observed in the TURP + TURBN group and the three indices were (6.2 +/- 3.0), (22.7 +/- 3.1) ml/s and (8.0 +/- 4.0) ml, respectively. No statistical difference (P > 0.05) was found in IPSS, Qmax and PVR among the three groups before the operation, but significant difference (P < 0.01) was observed after it. The curative effect was better in the TURP + TUIBN group than in the TURP, but was the best in the TURP + TURBN. CONCLUSIONS: TURP + TURBN, being more effective than TURP + TUIBN, should be used as the first option for the surgical treatment of small-size BPH. The key to the operation is to thoroughly remove not only the hyperplastic gland but also the pathological changes of the bladder neck.

Laparoscopy and laparoscopic ultrasonography in judging the resectability of pancreatic head cancer.

OBJECTIVE: To explore the clinical value of laparoscopy and laparoscopic ultrasonography (LUS) in judging the resectability of pancreatic head cancer. METHODS: LUS was employed as a prospective diagnosis of tumor staging before exploratory laparotomy in 22 patients diagnosed with pancreatic head cancer to identify whether the liver and peritoneum had small metastases or local invasion to the portal vein, superior mesenteric vessel, aorta, inferior vena cava. RESULTS: In the 22 patients receiving laparoscopy and LUS, we found peritoneal or surface liver metastases (3 patients), hepatic parenchyma metastases (1), and pancreatitis proved by biopsy under ultrasound guidance (1). Laparotomy was avoided in these 5 patients. Of the remaining 17 patients, 8 patients, including 2 patients with portal vein emboli due to tumor metastases had hypertrophic lymph nodes or tumor invasion of local vessels in the peritoneal cavity, retroperitoneum, and omentum and the other 9 patients had resectable tumors shown by LUS. The 17 patients were subjected to exploratory laparotomy, and pancreaticoduodenectomy was successful in 8 patients. CONCLUSIONS: Laparoscopy and LUS can be used to precisely estimate the possibility of resection of pancreatic head cancer, and prevent unnecessary exploratory laparotomy and its complications. It can be used as a routine examination before exploratory laparotomy.

Deletion of the telomerase reverse transcriptase gene and haploinsufficiency of telomere maintenance in Cri du chat syndrome.

Cri du chat syndrome (CdCS) results from loss of the distal portion of chromosome 5p, where the telomerase reverse transcriptase (hTERT) gene is localized (5p15.33). hTERT is the rate-limiting component for telomerase activity that is essential for telomere-length maintenance and sustained cell proliferation. Here, we show that a concomitant deletion of the hTERT allele occurs in all 10 patients with CdCS whom we examined. Induction of hTERT mRNA in proliferating lymphocytes derived from five of seven patients was lower than that in unaffected control individuals (P<.05). The patient lymphocytes exhibited shorter telomeres than age-matched unaffected individuals (P<.0001). A reduction in replicative life span and a high rate of chromosome fusions were observed in cultured patient fibroblasts. Reconstitution of telomerase activity by ectopic expression of hTERT extended the telomere length, increased the population doublings, and prevented the end-to-end fusion of chromosomes. We conclude that hTERT is limiting and haploinsufficient for telomere maintenance in humans in vivo. Accordingly, the hTERT deletion may be one genetic element contributing to the phenotypic changes in CdCS.