Circulating miR-31-5p may be a potential diagnostic biomarker in nasopharyngeal carcinoma.

Aberrant expression of miR-31-5p has been detected in various cancers and plays a significant role in tumorigenesis. Low miR-31-5p expression was present in nasopharyngeal carcinoma (NPC) tissues and cell lines and acted as a tumor suppressive miRNA. Currently, circulating miRNAs are emerging as novel biomarkers for the early diagnosis of cancers using a non-invasive method. However, circulating miR-31-5p has rarely been reported in NPC. Therefore, the aim of this study was to explore peripheral blood miR-31-5p levels as a noninvasive biomarker and evaluate its clinical value for the early diagnosis of patients with NPC. A total of 110 participants were recruited, including 55 NPC patients and 55 healthy controls. Peripheral blood samples were collected from these participants, and total RNA was extracted to quantify the relative expression of miR-31-5p by RT-qPCR. We found a significantly lower expression of miR-31-5p in the NPC patients than in the healthy controls. Furthermore, low expression of miR-31-5p was highly correlated with tumor-node-metastasis (TNM) stage (I+II vs III+IV, p=0.001), T classification (T1 vs T2+T3+T4, p=0.036) and local lymph node metastasis (N1-N3 vs N0, p=0.002), but not distant metastasis (p=0.288). Moreover, miR-31-5p showed a moderate diagnostic performance (AUC=0.866, sensitivity = 0.782, specificity = 0.818). Thus, we concluded that circulating miR-31-5p can be a potentially novel and non-invasive biomarker for the early diagnosis of NPC and an attractive therapeutic target in NPC patients.

Inhibition of the multidrug and toxin extrusion (MATE) transporter by pyrimethamine increases the plasma concentration of metformin but does not increase antihyperglycaemic activity in humans.

We hypothesized that the pharmacodynamic (PD) characteristics of metformin would change with inhibition of the multidrug and toxin extrusion (MATE) transporter, which mediates renal elimination of metformin. Twenty healthy male subjects received two doses (750/500 mg) of metformin, with and without 50 mg of pyrimethamine (a potent MATE inhibitor), with 1 week of washout in between each dose. The PD characteristics of metformin were assessed using oral glucose tolerance tests (OGTTs) before and after the metformin dose. Metformin concentrations in plasma and urine were determined using liquid chromatography-electrospray ionization-tandem mass spectrometry. When metformin was co-administered with pyrimethamine, its area under the concentration-time curve from 0 to 12 h was 2.58-fold greater (p < 0.05), whereas the antihyperglycaemic effects of metformin were decreased. The mean differences (90% confidence interval) in mean and maximum serum glucose concentrations and in 2-h-post-OGTT serum glucose concentration were -0.6 (-1, -0.2), -0.9 (-1.6, -0.3) and -0.5 (-1.1, 0.1) mmol/l, respectively. These findings indicate that the response to metformin is not only related to the plasma exposure of metformin but is also related to other factors, such as inhibition of uptake transporters and the gastrointestinal-based pharmacology of metformin.

Epidemiological investigation of MERS-CoV spread in a single hospital in South Korea, May to June 2015.

In this report, we describe 37 MERS-CoV infection cases (1 primary, 25 secondary, 11 tertiary cases) in a single hospital in South Korea. The median incubation period was six days (95% CI: 4­7 days) and the duration between suspected symptom onset and laboratory confirmation was 6.5 days (95% CI: 4­9). While incubation period was two days longer, the duration from suspected symptom onset to confirmation was shorter in tertiary compared with secondary infections.

Targeted disruption of the murine erythroid band 3 gene results in spherocytosis and severe haemolytic anaemia despite a normal membrane skeleton.

Band 3 is the most abundant integral protein of the red blood cell membrane. It performs two critical biological functions: maintaining ionic homeostasis, by transporting Cl- and HCO3-ions, and providing mechanical stability to the erythroid membrane. Erythroid band 3 (AE1) is one of three anion exchangers that are encoded by separate genes. The AE1 gene is transcribed by two promoters: the upstream promoter produces erythroid band 3, whereas the downstream promoter initiates transcription of the band 3 isoform in kidney. To assess the biological consequences of band 3 deficiency, we have selectively inactivated erythroid but not kidney band 3 by gene targeting in mice. Although no death in utero occurred, the majority of homozygous mice die within two weeks after birth. The erythroid band 3 null mice show retarded growth, spherocytic red blood cell morphology and severe haemolytic anaemia. Remarkably, the band 3-/- red blood cells assembled normal membrane skeleton thus challenging the notion that the presence of band 3 is required for the stable biogenesis of membrane skeleton. The availability of band 3-/- mice offers a unique opportunity to investigate the role of erythroid band 3 in the regulation of membrane-skeletal interactions, anion transport and the invasion and growth of malaria parasite into red blood cells.

[Controversy and progress on whether to retain left colonic artery in radical resection of rectal cancer].

Japanese Society for Cancer of the Colon and Rectum (JSCCR) guideline 2019 recommended that lymph node dissection for advanced rectal cancer should include the lymphatic adipose tissue at the root of the inferior mesenteric vessels, but the ligation site of the inferior mesenteric artery (IMA) was not determined, and the NCCN guideline did not indicate clearly whether to retain the left colonic artery (LCA). Controversy over whether to retain LCA is no more than whether it can reduce the incidence of anastomotic complications or postoperative functional damage without affecting the patients' oncological outcome. Focusing on the above problems, this paper reviews the latest research progress. In conclusion, it is believed that the advantages of retaining LCA are supported by most studies, which can improve the blood supply of the proximal anastomosis, and technically can achieve the same range of lymph node dissection as IMA high ligation. However, whether it affects the survival of patients, reduces the incidence of anastomotic leakage, and improves the quality of life of patients, more high-quality evidence-based medical evidence is still needed.

Enhanced wound healing using a 3D printed VEGF-mimicking peptide incorporated hydrogel patch in a pig model.

There is a need for effective wound healing through rapid wound closure, reduction of scar formation, and acceleration of angiogenesis. Hydrogel is widely used in tissue engineering, but it is not an ideal solution because of its low vascularization capability and poor mechanical properties. In this study, gelatin methacrylate (GelMA) was tested as a viable option with tunable physical properties. GelMA hydrogel incorporating a vascular endothelial growth factor (VEGF) mimicking peptide was successfully printed using a three-dimensional (3D) bio-printer owing to the shear-thinning properties of hydrogel inks. The 3D structure of the hydrogel patch had high porosity and water absorption properties. Furthermore, the bioactive characterization was confirmed by cell culture with mouse fibroblasts cell lines (NIH 3T3) and human umbilical vein endothelial cells. VEGF peptide, which is slowly released from hydrogel patches, can promote cell viability, proliferation, and tubular structure formation. In addition, a pig skin wound model was used to evaluate the wound-healing efficacy of GelMA-VEGF hydrogel patches; the results suggest that the GelMA-VEGF hydrogel patch can be used for wound dressing.

[Association between the expression of MMP1 gene and prognosis in head and neck squamous cell carcinoma].

Objective:The aim of this study is to investigate the expression of MMP1 and prognosis in patients with head and neck squamous cell carcinoma (HNSCC), and to identify the potential mechanism of MMP1 in HNSCC. Method:The RNA sequencing data and related clinical data of HNSCC were downloaded from the TCGA public database. The MMP1 gene expression data and corresponding clinical information in the samples were retrospectively analyzed; The data of gene microarray were used to verify the correlation between MMP1 gene and HNSCC. The disease free survival and overall survival of HNSCC were also analyzed; Gene set enrichment analysis was conducted to identify the potential mechanism of MMP1 in HNSCC. Result:Among the 332 HNSCC patients, the expression of MMP1 was significantly associated with lymphatic invasion and tumor grade (P<0.01). The higher the expression level of MMP1 was, the more susceptible the patient was to lymph node metastasis. The data confirmed that the expression of MMP1 in HNSCC was significantly higher than that in normal mucosa (P<0.05); HNSCC of patient in MMP1 high expression group proved to have worse disease free survival and overall survival than in MMP1 low expression group (P<0.05); Gene enrichment analysis indicates that the high expression of MMP1 gene might influence the biological process of tumor though epithelial mesenchymal transition, TGF-ß signaling pathway, hypoxia, angiogenesis, Noth signaling pathway, and up-regulation of KRAS gene signaling pathway. Conclusion:The high expression of MMP1 was related with occurrence and development of HNSCC, which can be used as an independent risk factor and has a great clinical significance.

[The efficacy comparation of adenoidectomy with acupuncture and tympanonstomy in children secretory otitis media].

Objective:This study aims to the comparative study of AT+A (adenoidectomy with acupuncture) and AT+T (adenoidectomy with tympanonstomy tube) to monitor and compare the therapeutic effect and prognosis of secretory otitis media in children. The study make a summary and give the clinical suggestions as well.Method:We collected and analyzed 280 outpatients of children secretory otitis media from March 2015 to March 2016.Among them,172 cases took the adenoidectomy with acupuncture and 108 cases took the adenoidectomy with tympanonstomy tube. This research used the therapeutic effect indicators,middle ear effusion time and one year follow-up to evaluate the pros and cons of two surgery methods in different areas.Result:The patients of both groups had relatively good therapeutic effect which promoted with time. There were no significant difference between AT+A and AT+T in tympanic membrane. While AT+T group acted better than AT+A group in pure tone average and tympanum figure. The middle ear effusion time of AT+T group was significantly shorter than AT+A group. In one year follow-up, there were no difference in hearing loss between two groups.But AT+T group performed better in recurrence rate, infection rate and total rate.Conclusion:Since the adenoidectomy with tympanonstomy tube method has a lot of advantages over adenoidectomy with acupuncture,it's better to use AT+T in severechildren secretory otitis media when situation is available.

Red cell membrane remodeling in sickle cell anemia. Sequestration of membrane lipids and proteins in Heinz bodies.

In red cells from patients with sickle cell anemia, hemoglobin S denatures and forms Heinz bodies. Binding of Heinz bodies to the inner surface of the sickle cell membrane promotes clustering and colocalization of the membrane protein band 3, outer surface-bound autologous IgG and, to some extent, the membrane proteins glycophorin and ankyrin. Loss of transbilayer lipid asymmetry is also found in certain populations of sickle red cells. The lateral distribution of sickle cell membrane lipids has not been examined, however. In this report, we examine by fluorescence microscopy the incorporation and distribution of the fluorescent phospholipid analogues 7-nitro-2,1,3-benzoxadiazol-4-yl (NBD)-phosphatidylserine and NBD-phosphatidylcholine in sickle red cells. Both phospholipid analogues are observed to accumulate prominently at sites of Heinz bodies. Accumulation at sites of Heinz bodies is also shown by 1,'1-dihexadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate, a fluorescent lipid analogue that readily crosses membranes, but not by fluorescein-phosphatidylethanolamine, an analogue that is localized to the outer leaflet of the membrane. Double labeling and confocal microscopy techniques show that NBD-lipids, band 3 protein, protein 4.1, ankyrin, and spectrin are all sequestered within sickle red cells and colocalized at sites of Heinz bodies. We propose that Heinz bodies provide a hydrophobic surface on which sickle red cell membrane lipids and proteins are sequestered.

Molecular basis of spectrin deficiency in beta spectrin Durham. A deletion within beta spectrin adjacent to the ankyrin-binding site precludes spectrin attachment to the membrane in hereditary spherocytosis.

We describe a spectrin variant characterized by a truncated beta chain and associated with hereditary spherocytosis. The clinical phenotype consists of a moderate hemolytic anemia with striking spherocytosis and mild spiculation of the red cells. We describe the biochemical characteristics of this truncated protein which constitutes only 10% of the total beta spectrin present on the membrane, resulting in spectrin deficiency. Analysis of reticulocyte cDNA revealed the deletion of exons 22 and 23. We show, using Southern blot analysis, that this truncation results from a 4.6-kb genomic deletion. To elucidate the basis for the decreased amount of the truncated protein on the membrane and the overall spectrin deficiency, we show that (a) the mutated gene is efficiently transcribed and its mRNA abundant in reticulocytes, (b) the mutant protein is normally synthesized in erythroid progenitor cells, (c) the stability of the mutant protein in the cytoplasm of erythroblasts parallels that of the normal beta spectrin, and (d) the abnormal protein is inefficiently incorporated into the membrane of erythroblasts. We conclude that the truncation within the beta spectrin leads to inefficient incorporation of the mutant protein into the skeleton despite its normal synthesis and stability. We postulate that this misincorporation results from conformational changes of the beta spectrin subunit affecting the binding of the abnormal heterodimer to ankyrin, and we provide evidence based on binding assays of recombinant synthetic peptides to inside-out-vesicles to support this model.

Duplication of 10 nucleotides in the erythroid band 3 (AE1) gene in a kindred with hereditary spherocytosis and band 3 protein deficiency (band 3PRAGUE).

We describe a duplication of 10 nucleotides (2,455-2,464) in the band 3 gene in a kindred with autosomal dominant hereditary spherocytosis and a partial deficiency of the band 3 protein that is reflected by decreased rate of transmembrane sulfate flux and decreased density of intramembrane particles. The mutant allele potentially encodes an abnormal band 3 protein with a 3.5-kD COOH-terminal truncation; however, we did not detect the mutant protein in the membrane of mature red blood cells. Since the mRNA levels for the mutant and normal alleles are similar and since the band 3 content is the same in the light and dense red cell fractions, we conclude that the mutant band 3 is either not inserted into the plasma membrane or lost from the membrane prior to the release of red blood cells into circulation. We further show that the decrease in band 3 content principally involves the dimeric laterally and rotationally mobile fraction of the band 3 protein, while the laterally immobile and rotationally restricted band 3 fraction is left essentially intact. We propose that the decreased density of intramembrane particles decreases the stability of the membrane lipid bilayer and causes release of lipid microvesicles that leads to surface area deficiency and spherocytosis.

The effect of beta-cyclodextrin complexation on the bioavailability and hepatotoxicity of clotrimazole.

Clotrimazole, a poorly water-soluble antimycotic agent, is a promising therapeutic agent for various diseases including cancer and sickle cell anemia. The oral bioavailability and hepatic toxicity of clotrimazole were compared with its beta-cyclodextrin inclusion form which was prepared by the spray-drying method. The inclusion complex gave significantly higher initial plasma concentrations, Cmax and AUC than did clotrimazole alone, indicating that the drug from the inclusion compound could be more easily absorbed in rats. Furthermore, mice treated with the inclusion compound showed significantly higher GOT/GPT values compared to clotrimazole alone. The inclusion compound also induced hypertrophy of hepatic cells by fat accumulation and disappearance of hepatic sinusoids, indications of pathological changes of liver, suggesting that the inclusion compound could induce more severe tissue damage in the liver than clotrimazole alone. Thus, hepatotoxicity of clotrimazole seems to be correlated with the enhanced oral bioavailability by inclusion complexation. Our results suggest that, in the development of a novel oral product, appearance or enhancement of hepatic toxicity must be considered along with oral bioavailability.

MiR-214 negatively regulates proliferation and WNT/ß-catenin signaling in breast cancer.

OBJECTIVE: To analyze the function and mechanism of miR-214 in regulating breast cancer cell proliferation. MATERIALS AND METHODS: MiR-214 expression level was measured by quantitative reverse transcription Polymerase Chain Reaction (qRT-PCR). The protein level of ß-catenin, PCNA and WNT targets (cyclin D1 and c-Myc) was evaluated by Western blot analysis. The effects of miR-214 on cell proliferation and cisplatin sensitivity were assessed by Cell Counting Kit-8 (CCK-8) assay and/or 5-bromo-2'-deoxyuridine (BrdU) assay. The effect of miR-214 on ß-catenin and WNT signaling activity was tested by luciferase reporter assay. RESULTS: MiR-214 was significantly downregulated in breast cancer tissues and was inversely correlated with ß-catenin expression. Forced expression of miR-214 in breast cancer cell line MCF-7 led to a decrease in cell proliferation and an increase in cisplatin sensitivity. Moreover, forced expression of miR-214 decreased the activity of WNT luciferase reporter and the luciferase reporter containing the 3'-untranslated region (3'UTR) of ß-catenin, whereas antisense inhibitor of miR-214 showed an opposite effect. Finally, miR-214 decreased the expression of ß-catenin and multiple WNT target genes. CONCLUSIONS: MiR-214 is downregulated and serves as a novel tumor suppressor in breast cancer. Forced expression of miR-214 in breast cancer cells diminished cancer cell survival possibly through inhibiting WNT signaling by direct repression of ß-catenin.

The replication of canine herpesvirus (CHV) induces apoptosis in canine kidney cell line: short communication.

The alphaherpesvirus canine herpesvirus (CHV) was tested in order to determine whether or not it has apoptotic potential. We have demonstrated that lytic replication of CHV resulted in induction of apoptosis. This phenomenon was confirmed using different techniques including in situ TUNEL assay and DNA laddering. The apoptotic activity of CHV might influence the pathobiology of this virus.

Effects of Allopurinol and Apocynin on Renal Ischemia-Reperfusion Injury in Rats.

BACKGROUND: This study evaluated the effects of allopurinol (ALP), a xanthine oxidase inhibitor, and apocynin (APC), a NADPH oxidase inhibitor, administered alone or together, on kidney damage caused by renal ischemia-reperfusion (IR) in rats. METHODS: Thirty rats were randomly assigned to 5 groups. Group 1 was a sham group. Group 2 was the renal IR control group (30-min ischemia followed by 24-h reperfusion). In groups 3 and 4, ALP or APC, respectively, was administered 1 h before the ischemia. In group 5, ALP and APC were co-administered. Blood urea nitrogen (BUN) and serum creatinine (Cr), renal tissue malondialdehyde (MDA) and superoxide dismutase (SOD), and histological changes were evaluated. RESULTS: A significant increase in BUN and Cr level, and histological damage was seen in the IR control group, indicating renal injury. Elevated MDA and decreased SOD levels in the IR control group demonstrated that renal damage occurred through oxidative stress. Pretreatment with ALP or APC alone or together prevented IR-induced renal damage. However, there was no significant difference between treatment with a single drug and co-administration of ALP and APC. CONCLUSIONS: The use of ALP and/or APC before ischemia may be beneficial to ameliorate renal IR injury.

Inhibition by phenytoin of in vitro secretion of calcitonin from rat thyroid glands and cultured rat C cells.

Baby rat thyroid glands and cultured rat medullary carcinoma C cells were incubated acutely with phenytoin (38-100 microM), and the calcitonin (CT) secreted into the serum-free medium was measured by radioimmunoassay (RIA). Phenytoin did not alter CT release from glands or C cells incubated in 1 mM Ca, but, when Ca was raised to 1.75 or 2.5 mM, a marked inhibitory effect of phenytoin was apparent. The inhibitory effect could be negated by including 10 microM BAY-K-8644 in the medium. Inhibitory effects on CT release also were obtained with 100 microM trifluoperazine or 100 microM nitrendipine, and these inhibitory effects also were counteracted by 10 microM BAY-K-8644. The results show that clinically relevant amounts of phenytoin can inhibit CT release, perhaps by interfering with C-cell Ca channels or by inhibiting calmodulin-dependent processes.

Corticotropin-releasing hormone mediates the response to cold stress in the neonatal rat without compensatory enhancement of the peptide's gene expression.

A variety of stressors activate the hypothalamic-pituitary-adrenal axis, with secretion and compensatory enhanced synthesis of hypothalamic corticotropin-releasing hormone (CRH). Whether CRH is a major effector in the stress response of the neonatal rat and whether the peptide's gene expression is subsequently up-regulated are not fully understood. We studied the effect of cold-separation stress on plasma corticosterone (CORT) levels and CRH messenger RNA (CRH-mRNA) abundance in the paraventricular nucleus. Rats (4-16 days old) were subjected to maximal tolerated cold-separation. CORT and CRH-mRNA abundance were measured before and at several time points after stress. Cold-separation stress resulted in a significant plasma CORT increase in all age groups studied. This was abolished by the administration of an antiserum to CRH on both postnatal days 6 and 9. CRH-mRNA increased in rats aged 9 days or older, but not in 6-day-old rats, by 4 h after stress. These results suggest the presence of robust CRH-mediated adrenal responses to cold-separation stress in neonatal rats. Before postnatal day 9, however, the compensatory increase in CRH-mRNA abundance is minimal.

Plasmodium falciparum erythrocyte membrane protein 1 is anchored to the actin-spectrin junction and knob-associated histidine-rich protein in the erythrocyte skeleton.

A distinctive pathological feature of Plasmodium falciparum malaria is the endothelial attachment of erythrocytes infected with mature asexual-stage parasites in microvessels of the major organs. Electron-dense protrusions described as knobs are displayed on the surface of parasitized erythrocytes and act as attachment points in cytoadherence. Parasite-encoded knob-associated histidine-rich protein (KAHRP) is a major component of knobs found on the cytoplasmic side of the host cell membrane. P. falciparum erythrocyte membrane protein 1 (PfEMP1) is a family of parasite-encoded cytoadherence receptors localized to knobs on the surface of parasitized erythrocytes. Despite its high antigenic diversity, PfEMP1 has a remarkably conserved cytoplasmic domain. We demonstrate in this study that the cytoplasmic domain of PfEMP1 (VAR(CD)) binds to host spectrin and actin and to full-length KAHRP in vitro. Apparent dissociation constants determined for VAR(CD)/F-actin and VAR(CD)/KAHRP interactions are 44.9+/-6.4 and 10. 7+/-2.2 nM, respectively. Further, we provide evidence that KAHRP polypeptides self-associate in solution to form structures similar to knobs and show binding of self-associated KAHRP clusters to spectrin-actin-protein 4.1 complexes. Findings in this study suggest that PfEMP1 is localized to the knob in P. falciparum-infected erythrocytes by binding to the host spectrin-actin junction and to self-associated KAHRP through its conserved cytoplasmic domain.

Effects of acute and chronic administration of cocaine on striatal uptake, compartmentalization and release of [3H]dopamine.

The effects of acute and repetitive administration of cocaine were studied on several parameters associated with the uptake and release of [3H]dopamine ([3H]DA) in the striatum. It was found that repetitive administration of cocaine followed 7 days later by acute challenge with cocaine, produced an increase in the Vmax with no change in the affinity of the uptake carrier for either dopamine (DA) or cocaine. The intracellular compartmentalization of [3H]DA in synaptosomes was not altered by either acute or repeated treatment with cocaine. However, chronic administration of cocaine abolished the stimulatory effect that 1 microM amphetamine normally has on the efflux of [3H]DA from the fast pool in untreated synaptosomes. The K(+)-stimulated release of [3H]DA from slices of striatum was not affected by acute or chronically administered cocaine; however, chronically administered cocaine, plus acute challenge with cocaine potentiated the effect of amphetamine on the K(+)-induced release of [3H]DA. This was accompanied by a reduction of the effect of amphetamine on the spontaneous release of DA. In addition, chronically administered cocaine plus acute challenge with cocaine increased K(+)-stimulated release of [14C]acetylcholine [( 14C]ACh). These data suggest that repetitive administration of cocaine, in a regimen that elicits behavioral sensitization, alters the substrates through which amphetamine exerts its effects on the subcellular distribution and release of [3H]DA, and further, that challenge with cocaine of sensitized rats produces a compensatory increase in the uptake of [3H]DA that is correlated with increased depolarization-induced release of [14C]ACh.

Rac GTPase activity is essential for EGF-induced mitogenesis.

Rac, a member of the Rho family GTPases, has been implicated in the regulation of a wide range of biological processes including actin remodeling, cell transformation, G1 cell cycle progression, and gene expression. To determine whether Rac GTPase activity is required for epidermal growth factor-induced mitogenesis, Rat-2 stable cells expressing a dominant-negative Rac1 mutant, RacN17, were prepared. Exposure to EGF exhibited a significantly restricted growth response in Rat-2-RacN17 cells compared to Rat-2 parental cells, suggesting an essential role of Rac in EGF-induced mitogenesis. In contrast, addition of lysophosphatidic acid exerted the same level of growth in Rat-2 and Rat-2-RacN17 cells. To gain further evidence for the essential role of Rac in EGF-induced mitogenesis, we performed the microinjection experiment. EGF-induced DNA synthesis was significantly blocked by microinjection of recombinant RacN17 protein, and not control IgG. Our further study to analyze the downstream mediator of Rac in EGF-signaling to mitogenesis demonstrated that Rac-activated phospholipase A2 plays a critical role. Taken together, our results suggest that the "Rac and Rac-activated PLA2" cascade is one of the major mitogenic pathways induced by EGF.