Suppression of αvß6 downregulates P-glycoprotein and sensitizes multidrug-resistant breast cancer cells to anticancer drugs.

Multidrug resistance (MDR) in breast cancer treatment is the major cause leading to the failure of chemotherapy. P-glycoprotein (P-gp), the product of the human MDR1 gene, plays a key role in resistance to chemotherapy and confers cross-resistance to many structurally unrelated anticancer drugs. We have previously reported that integrin αvß6 plays a critical role in breast cancer invasion and metastasis. However, whether and how αvß6 is associated with P-gp and regulated by potential genetic mechanisms in breast cancer remains unclear. In the present study, we further investigated the reversal effect and underlying mechanisms of MDR in breast cancer. Two small interfering RNA constructs (pSUPER-ß6shRNAs) targeting two different regions of the ß6 gene have been designed to inhibit αvß6 expression by transfecting them into adriamycin-resistant MCF-7/ADR cell lines. Suppression of αvß6 dramatically downregulated the levels of MDR1 gene mRNA and P-gp. In particular, ß6shRNA-mediated silencing of αvß6 gene increased significantly the cellular accumulation of Rhodamine 123 and markedly decreased drug efflux ability, suggesting that ß6shRNAs indeed inhibit P-gp mediated drug efflux and effectively overcome drug resistance. In addition, inhibition of integrin αvß6 suppressed the expression of ERK1/2. Interestingly, our data demonstrate that suppression of integrin αvß6 caused significant downregulation of Bcl-2, Bcl-xL and upregulation of caspase 3, Bad, accompanied by increasing activity of cytochrome C. A possible connection between αvß6 and P-gp in drug resistance biology is suggested. Taken together, ß6shRNA could efficiently inhibit αvß6 and MDR1 expression in vitro and these findings may offer specifically useful means to reverse MDR in breast cancer therapy.

[Characteristics of acute myocardial infarction caused by spontaneous coronary artery dissection in young female patients].

Objective: To investigate the characteristics of acute myocardial infarction caused by spontaneous coronary artery dissection(SCAD) in young female patients. Methods: In this casecontrolstudy,127 young(≤55 years) female patients with acute myocardial infarction onset within 1 week in Nanjing first hospital, Xuzhou central hospital, affiliated hospital of Xuzhou medical university, and Lianyungang first people's hospital were enrolled between January 2013 and February 2017,and the clinical data were retrospectively analyzed. According to their clinical manifestations and coronary angiography(CAG) results,the patients were divided into coronary atherosclerosis disease(CAD) group(CAG evidenced atherosclerosis, n=83) and SCAD group(CAG detected coronary artery dissection,n=44).The SCAD patients were subdivided into definite group (the results affirmed from intravenous ultrasound or optical coherence tomography, n=21) and probable group (the CAG results highly confirmed to characteristics of SCAD,but no intravenous ultrasound or optical coherence tomography image affirmation,n=23). Then, according to the different treatment strategies, the SCAD patients were subdivided into conservative treatment group(treated with drugs,n=19) and interventional therapy group(treated with percutaneous coronary intervention,n=25). Results: (1)Compared to CAD group, patients in the SCAD group had less risk factors, such as hypertension history (25.0% (11/44) vs. 45.8% (38/83) , P=0.022) and diabetes history (6.8% (3/44) vs. 21.7% (18/83) , P=0.043),and had lower levels of fasting blood glucose (5.34(4.59,5.87) mmol/L vs. 7.12(5.18,8.60)mmol/L, P=0.001),total cholesterol((3.94±1.14) mmol/L vs. (4.91±1.50) mmol/L, P=0.001),triglyceride(1.42 (0.91,1.64) mmol/L vs. 1.89 (1.23,2.45) mmol/L, P=0.005),and low density lipoprotein cholesterol ((2.24±0.91) mmol/L vs. (2.94±1.16) mmol/L, P=0.001),CAG results showed that patients in the SCAD group had more single vessel lesion (88.6% (39/44) vs. 39.8% (33/83) , P=0.001), and their target lesion stenosis was less severe ( (79.2±22.4) % vs. (91.5±12.1) %, P=0.001). (2) The clinical risk factors such as hypertension history, diabetes history, smoking history, family history of cardiology disease, fasting blood glucose,total cholesterol,triglyceride and low density lipoprotein cholesterol were similar between definite group and probable group (all P>0.05). CAG results showed that prevalence of single vessel lesion (100% (21/21) vs. 78.3% (18/23) , P=0.050) and percent of target lesion stenosis ( (76.9±20.6) % vs. (81.2±24.1) %, P=0.529) were similar between definite group and probable group.(3)There were no significant difference in single vessel(84.0% (21/25) vs. 94.7% (18/19) , P=0.370), target lesion stenosis(85.0(70.0,100.0)% vs. 75.0(50.0,90.0)%, P=0.186),and survival rates in hospital(96.0% (24/25) vs. 100% (19/19) , P=1.000) between interventional therapy group and conservative treatment group. Conclusions: Prevalence of SCAD is highin young female patients with acute myocardial infarction. Acute myocardial infarction patients with less risk factors of CAD and with CAG showing smooth lesion of narrowing segment and normal finding in the other vessels, are more likely to be diagnosed with SCAD.Acute myocardial infarction patients caused by SCAD have high survival rate either receiving percutaneous coronary intervention or drug treatment.

[Role of bone marrow mesenchymal stem cells with CTLA4Ig and CD40LIg gene modification in rejection reaction after liver transplantation].

Objective: To investigate the role of bone marrow mesenchymal stem cells (BMSCs) with CTLA4Ig and CD40LIg gene modification in rejection reaction after liver transplantation in rats and possible mechanisms. Methods: The modified Kamada's two-cuff technique was used to establish a Lewis-BN rat model of orthotopic liver transplantation, and a total of 75 rats were randomly divided into groups A, B, C, D, and E, with 15 rats in each group. The rats in group A (control group) were given infusion of isotonic saline via the portal vein during liver transplantation, those in group B (BMSC group) were given infusion of BMSCs via the portal vein during liver transplantation, those in group C (BMSCs with CTLA4Ig gene modification) were given infusion of BMSCs carrying the CTLA4Ig gene via the portal vein during liver transplantation, those in group D (BMSCs with CD40LIg gene modification) were given infusion of BMSCs carrying the CD40LIg gene via the portal vein during liver transplantation, and those in group E (BMSCs with CTLA4Ig and CD40LIg gene modification) were given infusion of BMSCs carrying CTLA4Ig and CD40LIg gene modification via the portal vein during liver transplantation. Postoperative survival and change in liver function were observed. HE staining was used to observe the pathomorphological changes of the graft liver, and ELISA was used to measure the levels of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-10 (IL-10), and interferon-γ (IFN-γ) in peripheral blood. A one-way analysis of variance was used for comparison of means of multiple samples, and the Kaplan-Meier survival curve analysis was used for comparison of survival rates between multiple groups. Results: Group E had a significantly longer survival time after surgery than groups A, B, C, and D (P < 0.05), groups C and D had a significantly longer survival time than groups A and B (P < 0.05), and there was no significant difference between groups C and D (P > 0.05). On day 10 after surgery, group A had significantly higher levels of alanine aminotransferase and total bilirubin than the other four groups (P < 0.05). HE staining showed severe rejection reaction in group A, moderate rejection reaction in group B, and mild rejection reaction in groups C and D; pathological examination showed no marked rejection reaction in group E. Group A had significant increases in the levels of IL-2 and IFN-γ and significant reductions in the levels of IL-4 and IL-10 after surgery compared with the other four groups (all P < 0.05). Conclusion: Infusion of BMSCs with modification of both CTLA4Ig and CD40LIg genes can significantly inhibit acute rejection reaction after liver transplantation in rats and effectively prolong the survival time of the graft liver, with a better effect than infusion of BMSCs alone or BMSCs with modification of CTLA4Ig or CD40LIg gene.

Liver abscesses in adult patients with and without diabetes mellitus: an analysis of the clinical characteristics, features of the causative pathogens, outcomes and predictors of fatality: a report based on a large population, retrospective study in China.

In China, there are four types of liver abscesses (LAs) that meet the clinical criteria. Pyogenic liver abscesses (PLAs) and amoebic liver abscesses (ALAs) are two of the most common types of abscesses, followed by fungal liver abscesses (FLAs) and hydatid secondary liver abscesses (HsLAs). Diabetes mellitus (DM) is associated with the development of PLAs. However, there is a lack of population-based studies that have evaluated the underlying relationship between LAs (mainly PLAs and FLAs) and DM. We conducted a retrospective study based on a large population to identify the potential differences and factors that affect the mortality of PLA patients in DM and non-DM groups. Our results revealed that the prevalence of DM is 44.3% (158/357) in PLA patients and 35.3% (18/51) in FLA patients. Compared with the non-DM patients, statistically significant differences were found in DM patients according to symptomatology, clinical manifestations, laboratory findings, microbiological characteristics, antimicrobial resistance, clinical treatments and outcomes in relation to mortality. In addition, the status of antibiotic resistance to E. coli and K. pneumoniae, which were isolated from the patient samples, is severe in the area in which the study was conducted. Regarding the treatment of PLAs, our study indicated that broad-spectrum antimicrobial therapy and drug combinations should be recommended and initiated before the pathogens are cultured and identified. In the clinic, therapies that combine percutaneous drainage with antibiotics and surgery with antibiotics are the two most useful strategies for treating an LA. These two combined treatments resulted in satisfactory cure rates. In the DM and non-DM groups, the cure rates for percutaneous drainage with antibiotics were 90.3% and 92.0%, respectively, and the cure rates for surgery with antibiotics were 93.9% and 95.2%, respectively.

Death the Fas way: regulation and pathophysiology of CD95 and its ligand.

Apoptotic cell death mediated by the members of the tumor necrosis factor receptor family is an essential process involved in the regulation of cellular homeostasis during development, differentiation, and pathophysiological conditions. Among the cell death receptors comprising the tumor necrosis factor receptor superfamily, CD95/APO-1 (Fas) is the best characterized. The specific interaction of Fas with its cognate ligand, Fas ligand (FasL), elicits the activation of a death-inducing caspase (cysteine aspartic acid proteases) cascade, occurring in a transcription-independent manner. Caspase activation executes the apoptosis process by cleaving various intracellular substrates, leading to genomic DNA fragmentation, cell membrane blebbing, and the exposure of phagocytosis signaling molecules on the cell surface. Recent studies have shown that the Fas/FasL pathway plays an important role in regulating the life and death of the immune system through activation-induced cell death. In addition, these molecules have been implicated in aging, human immunodeficiency virus infection, drug abuse, stress, and cancer development. In this review, we will focus on the mechanisms that regulate Fas and FasL expression, and how their deregulation leads to diseases.

Suppression of angiotensin II stimulated responses in aortic vascular smooth muscle cells of experimental cirrhotic rats.

Functional responses to angiotensin II (AT-II) were determined in aortic vascular smooth muscle cells (VSMCs) from experimental cirrhotic rats. Our data showed that AT-II-stimulated extracellular acidification rate (ECAR), which was measured by Cytosensor microphysiometry, was significantly reduced in the aortic VSMCs from the cirrhotic rats as compared to those from the control animals. The ability of AT-II to promote formation of inositol phosphates, the second messenger produced by the activation of Gq-coupled receptors, was also considerably suppressed in the cirrhotic VSMCs. Furthermore, the maximal p42/44 MAPK phosphorylation stimulated by AT-II was significantly reduced in the cirrhotic VSMCs in contrast to that in the normal VSMCs. Taken together, our data clearly demonstrated that the functional responses to AT-II was severely suppressed in aortic VSMCs in cirrhosis, indicating the impairment of general Gq-coupled receptor signaling and subsequent biological function in the cirrhotic VSMCs.

Differential neurotoxicity of etorphine-like opiates: lack of correlation with their ability to activate opiate receptors.

The present study was undertaken to compare the neurotoxic effects of three etorphine-like opiates (etorphine, dihydroetorphine, and another derivative of oripavine) and heroin with their ability to activate opiate receptors in human neuroblastoma cell line SK-N-SH as well as in two other neuronal cell lines. Neurotoxicity was measured by using [3H]-thymidine incorporation analysis, cell viability measurement and Cytosensor microphysiometry. It was found that, in spite of the very similar molecular structures of these opiates, they displayed significant differences in cytotoxicity, with etorphine and another derivative of oripavine possessing high potency but dihydroetorphine and heroin little effect. However, neurotoxic potency of the opiates was not directly correlated to their ability to activate opioid receptors, as determined by [35S]-guanylyl-5'-O-(gamma-tho)-triphosphate binding assay. These findings provide clear evidence of differential neurotoxicity of etorphine-like opiates, and suggest that the neurotoxicity is not closely related to the molecular configuration required as opioid receptor agonist but is probably associated with the presence of a double bond in the structure.

Antisense oligonucleotide to insulin-like growth factor II induces apoptosis in human ovarian cancer AO cell line.

The effects of antisense oligonucleotide to insulin-like growth factor II (IGFII) to induce apoptosis in human ovarian cancer cells were evaluated. antiproliferation effects of antisense to IGFII in ovarian cancer AO cells were determined by 3H-thymidine incorporation. Apoptosis of the IGFII antisense-treated cells was quantitated by both nuclear condensation and flow cytometry after cells were stained with propidium iodide. IGFII antisense (4.5 microM) treatment of 48 h maximally inhibited proliferation of AO cells. More than 25% of IGFII antisense-treated cells (4.5 microM for 24 h) had undergone apoptosis, whereas less than 3% of the cells were apoptotic in either IGFII sense-treated cells or untreated cells. Antisense oligonucleotide to IGFII significantly inhibited cell proliferation and induced apoptosis in human ovarian cancer AO cell. These data suggest that IGFII may be a potential target in treatment of ovarian cancer and antisense oligonucleotide to IGFII may serve as a therapeutic approach.

Etorphine inhibits cell growth and induces apoptosis in SK-N-SH cells: involvement of pertussis toxin-sensitive G proteins.

Opiates have been used extensively in the treatment of pain but with the severe side effect of addiction, which is believed to be related to opiates' direct (primary) or indirect (secondary) neurotoxicity. In this study, the effects of opioids on cell growth and apoptosis have been examined in human neuroblastoma cell line SK-N-SH. Etorphine, a wide-spectrum and potent agonist of opioid receptors, was found to significantly inhibit cell growth and to induce apoptosis. The inhibitory and apoptotic activities of etorphine followed a dose- and time-dependent manner. The more specific agonists of opioid receptors such as morphine, [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAGO), [D-Pen2, D-Pen5]-enkephalin (DPDPE), dynorphin A and nociceptin/orphanin FQ did not show similar toxic activities under the same conditions. In addition, the effects of etorphine could not be blocked by the opioid receptor antagonist naloxone, suggesting that the effects of etorphine might not be mediated by a classical opioid receptor. However, pretreatment of SK-N-SH cells with pertussis toxin (PTX) blocked the inhibition of cell growth and apoptosis induced by etorphine, indicating the involvement of PTX-sensitive G proteins in the processes. It was also shown that etorphine-induced apoptosis was prevented by actinomycin D (AD) and interleukin-1beta converting enzyme inhibitor I. Interestingly, etorphine was similarly potent to inhibit growth of pheochromocytoma (PC12) cells but less effective in SH-SY5Y neuroblastoma cells and C6 glioma cells. We propose that inhibition of cell growth and induction of apoptosis may be one mechanism of opioid neurotoxicity.

Retardation of pain development: a case of recovery from congenital insensitivity to pain.

Congenital analgesia is a rare genetic disorder. We report here that a 12-year-old boy was able to recover from congenital insensitivity to pain. Neurological examinations revealed that there was a 'stocking' distribution of pain decrement on the lower extremities under the patient's knee joints. Magnetic Resonance Imaging (MRI) of his brain showed gyrus thinning with sulcus widening at both sides of the parietal lobe. Southern blot hybridization probed with cDNAs of various opioid receptors did not detect any significant abnormality. Our results suggest that this rare case may not be genetically determined.

Progesterone induces apoptosis and up-regulation of p53 expression in human ovarian carcinoma cell lines.

BACKGROUND: Progesterone (PROG) has been shown to reduce the risk of developing ovarian carcinoma in postmenopausal women who have undergone estrogen and progestogen replacement therapy, and it has been clinically used to treat some types of ovarian tumors. It is not yet clear whether or not the antitumor activity of progestogen is due to its ability to induce apoptosis in precarcinomatous and carcinomatous ovarian cells. The apoptosis-related genes p53, bcl-2, and c-myc have important roles in the regulation of programmed cell death, and thus may be involved in the process of the suspected PROG-induced apoptosis. METHODS: Antiproliferation effects of PROG on 3AO and AO ovarian carcinoma cells were determined by 3H-thymidine incorporation. Apoptosis of the PROG-treated cells was determined by DNA laddering analysis and was quantitated by both nuclear condensation and flow cytometry after cells were stained with propidium iodide. Cell cycle analysis was also performed by flow cytometry. The expression of p53, bcl-2, and c-myc after 72 hours of PROG treatment was detected by Northern blot analysis. RESULTS: In both 3AO and AO cell lines, cells proliferation was maximally inhibited by PROG after 72 hours of treatment at 10 microM concentration. Under the same conditions, more than 50% of PROG-treated cells had undergone apoptosis, whereas less than 3% of the cells were apoptotic in untreated cell cultures. After exposure to PROG for 72 hours, cells were arrested in the G1 phase of the cell cycle, and the levels of p53 mRNA were remarkably increased in both cell lines. No changes in expression of bcl-2 or c-myc were detected. CONCLUSIONS: PROG significantly inhibited cell proliferation and induced apoptosis in both of the ovarian carcinoma cell lines tested in this study. PROG treatment markedly up-regulated p53 expression in these cells, indicating involvement of p53 in PROG-induced apoptosis.

[Diagnosis and treatment of cysticercosis of cerebral ventricles].

31 patients with cysticercosis of cerebral ventricles verified by operation or pathological investigation were reported. All patients were between 7 and 64 years of age and 14 were females. All had a single cyst. Since 29 patients (94%) were without a history of intestinal taeniasis, it was proposed that most patients of cysticercosis of cerebral ventricles were caused by hetero-infection and the entrance of Cysticercus into brain ventricle was through choroid plexus along the cerebrospinal fluid. This is probably the reason why it occurs mostly in the 4th ventricle. The clinical manifestation of cysticercosis of cerebral ventricles were paroxysmal headache and vomiting caused by increased intracranial pressure. Ventriculography and CT scanning have considerable diagnostic value. Removal of Cysticercus by surgical operation is successful (Figs. 1-8).

[Chemistry and antitumor activity of hainanensine analogues].

A synthetic analogue of hainanensine 2a was further modified in order to improve its antitumor activity. Six compounds 3-8, were obtained, and among these compounds 8 showed significant activity. The diastereomer (+/-)8B was resolved by (+) tartaric acid and (-)benzoyl-tartaric acid to the corresponding enantiomers (-)8B and (+)8B, respectively and their optical purities were determined by isotope dilution method as 93% and 96%. The antitumor activity of the newly obtained compounds were tested in vitro, and (-)8B was found to have an IC50 of 1.9 mol/L, eighteen times higher than the original compound 2a.