Phosphorylated PKM2 regulates endothelium-dependent vasodilation in diabetes / 中南大学学报(医学版)

OBJECTIVES@#Endothelium-dependent vasodilation dysfunction is the pathological basis of diabetic macroangiopathy. The utilization and adaptation of endothelial cells to high glucose determine the functional status of endothelial cells. Glycolysis pathway is the major energy source for endothelial cells. Abnormal glycolysis plays an important role in endothelium-dependent vasodilation dysfunction induced by high glucose. Pyruvate kinase isozyme type M2 (PKM2) is one of key enzymes in glycolysis pathway, phosphorylation of PKM2 can reduce the activity of pyruvate kinase and affect the glycolysis process of glucose. TEPP-46 can stabilize PKM2 in its tetramer form, reducing its dimer formation and phosphorylation. Using TEPP-46 as a tool drug to inhibit PKM2 phosphorylation, this study aims to explore the impact and potential mechanism of phosphorylated PKM2 (p-PKM2) on endothelial dependent vasodilation function in high glucose, and to provide a theoretical basis for finding new intervention targets for diabetic macroangiopathy.@*METHODS@#The mice were divided into 3 groups: a wild-type (WT) group (a control group, C57BL/6 mice) and a db/db group (a diabetic group, db/db mice), which were treated with the sodium carboxymethyl cellulose solution (solvent) by gavage once a day, and a TEPP-46 group (a treatment group, db/db mice+TEPP-46), which was gavaged with TEPP-46 (30 mg/kg) and sodium carboxymethyl cellulose solution once a day. After 12 weeks of treatment, the levels of p-PKM2 and PKM2 protein in thoracic aortas, plasma nitric oxide (NO) level and endothelium-dependent vasodilation function of thoracic aortas were detected. High glucose (30 mmol/L) with or without TEPP-46 (10 μmol/L), mannitol incubating human umbilical vein endothelial cells (HUVECs) for 72 hours, respectively. The level of NO in supernatant, the content of NO in cells, and the levels of p-PKM2 and PKM2 protein were detected. Finally, the effect of TEPP-46 on endothelial nitric oxide synthase (eNOS) phosphorylation was detected at the cellular and animal levels.@*RESULTS@#Compared with the control group, the levels of p-PKM2 in thoracic aortas of the diabetic group increased (P<0.05). The responsiveness of thoracic aortas in the diabetic group to acetylcholine (ACh) was 47% lower than that in the control group (P<0.05), and that in TEPP-46 treatment group was 28% higher than that in the diabetic group (P<0.05), while there was no statistically significant difference in the responsiveness of thoracic aortas to sodium nitroprusside (SNP). Compared with the control group, the plasma NO level of mice decreased in the diabetic group, while compared with the diabetic group, the phosphorylation of PKM2 in thoracic aortas decreased and the plasma NO level increased in the TEPP-46 group (both P<0.05). High glucose instead of mannitol induced the increase of PKM2 phosphorylation in HUVECs and reduced the level of NO in supernatant (both P<0.05). HUVECs incubated with TEPP-46 and high glucose reversed the reduction of NO production and secretion induced by high glucose while inhibiting PKM2 phosphorylation (both P<0.05). At the cellular and animal levels, TEPP-46 reversed the decrease of eNOS (ser1177) phosphorylation induced by high glucose (both P<0.05).@*CONCLUSIONS@#p-PKM2 may be involved in the process of endothelium-dependent vasodilation dysfunction in Type 2 diabetes by inhibiting p-eNOS (ser1177)/NO pathway.

Purification and characterization of a thermostable alkaline cellulase produced by Bacillus licheniformis 380 isolated from compost

ABSTRACT During composting processes, the degradation of organic waste is accomplished and driven by a succession of microbial populations exhibiting a broad range of functional competencies. A total of 183 bacteria, isolated from a composting process, were evaluated for cellulase activity at different temperatures (37, 50, 60, and 70°C) and pH values. Out of the 22 isolates that showed activity, isolate 380 showed the highest cellulase activity. Its ability to produce cellulase was evaluated in culture medium supplemented with carboxymethyl cellulose, microcrystalline cellulose, wheat straw, and rice husk. The culture medium supplemented with carboxymethyl cellulose induced higher enzyme activity after 6 hours of incubation (0.12 UEA mL-1 min-1). For wheat straw and rice husk, the results were 0.08 UEA mL-1 min-1 for both, while for microcrystalline cellulose, 0.04 UEA mL-1 min-1 were observed. The highest carboxymethyl cellulase activity was observed at 60°C (0.14 UEA mL-1 min-1) for both crude and partially purified enzyme after 30 and 120 min of incubation, respectively. Alkalinization of the medium was observed during cultivation in all substrates. The cellulase had a molecular mass of 20 kDa determined by SDS-Page. Isolate 380 was identified as Bacillus licheniformis. This work provides a basis for further studies on composting optimization.

Comparable bone healing capacity of different bone graft matrices in a rabbit segmental defect model

We compared the bone healing capacity of three different demineralized bone matrix (DBM) products applied using different carrier molecules (hyaluronic acid [HA] vs. carboxymethylcellulose [CMC]) or bone compositions (cortical bone vs. cortical bone and cancellous bone) in a rabbit segmental defect model. Overall, 15-mm segmental defects in the left and right radiuses were created in 36 New Zealand White rabbits and filled with HA-based demineralized cortical bone matrix (DBX), CMC-based demineralized cortical bone matrix (DB) or CMC-based demineralized cortical bone with cancellous bone (NDDB), and the wound area was evaluated at 4, 8, and 12 weeks post-implantation. DBX showed significantly lower radiopacity, bone volume fraction, and bone mineral density than DB and NDDB before implantation. However, bone healing score, bone volume fraction, bone mineral density, and residual bone area at 4, 8, and 12 weeks post-implantation revealed no significant differences in bone healing capacity. Overall, three DBM products with different carrier molecules or bone compositions showed similar bone healing capacity.

Standardization of serum free media for cryopreservation of RK13 cells.

Foetal calf serum present in the media used for cryopreservation was replaced by various synthetic polymer such as gelatin, glycerol, carboxymethyl cellulose and dimethyl sulphoxide at various concentration. Growth pattern of cells, % survival and karyological studies have been done in the present study. It was found that optimum concentration of carboxymethyl cellulose was 0.1% in combination with 10% glycerol and 10% DMSO. At this concentration percentage survival of cells was found maximum and karyotype was found normal without any abnormality in the chromosomes. It was concluded from the study that serum free media can be employed for the cryopreservation of these cells which are further used for production of tissue culture vaccines without causing any adverse affects.

Effect of sodium carboxymethylcellulose and methylprednisolone on the healing of jejunal anastomoses in rats

Sodium carboxymethylcellulose (SCMC) has been effective in reducing adhesion formation and corticosteroids reduce the inflammatory process. The objective of this study was to define the intraperitoneal (ip) effects of SCMC combined with intramuscular (im) methylprednisolone on peritoneal adhesion formation and on jejunal anastomosis healing in rats. Twenty Wistar rats (200-350 g) were divided into four groups (N = 5): groups I and III (controls) 5 and 21 days of treatment before sacrifice, respectively; groups II and IV (experimental groups) 5 and 21 days of treatment, respectively. SCMC (1 percent) was infused into the abdominal cavity and methylprednisolone (10 mg kg-1 day-1) was injected im daily from the day before surgery for animals of groups II and IV. All rats were submitted to a jejunal anastomosis. Sections of the anastomosis were prepared for routine histopathological analysis. The abdominal adhesion of group IV was less intense when compared with group III (P<0.0008). Anastomotic resistance was higher in groups II and IV when compared with groups I and III, respectively (P<0.05). There was no histological difference between groups I and II (exuberant granulation tissue on the serosal surface). Group III presented little peritoneal fibrinous tissue, with numerous thick collagen fibers. Group IV presented extensive although immature young fibrous tissue with rare thick collagen fibers. Sodium carboxymethylcellulose combined with corticosteroids seemed to diminish peritoneal adhesion but did not reduce anastomotic resistance

Esquema terapéutico en las úlceras de miembros inferiores: experiencia con la bota de Unna modificada / Therapeutic schedule in the lower limb ulcers: experience with modified Unna boot

Las úlceras de miembros inferiores por insuficiencia venosa son una patología frecuente en pacientes por sobre la cuarta década de la vida, incapacitante, refractaria a la terapia y causal de hospitalización prolongada. La Pasta de Unna ha sido uno de los tratamientos más utilizados desde los últimos 100 años. Mostramos en este trabajo los beneficios del uso de un esquema secuencial de la cicatrización completa o preparación para un ulterior injerto de dichas úlceras; esta experiencia se fundamenta en otras investigaciones publicadas con anterioridad. Realizamos un estudio prospectivo en 23 pacientes, de ambos sexos, adultos, con úlceras de miembros inferiores, de diferentes etiologías, durante el periodo comprendido entre enero de 1986 hasta octubre de 1998. Se utilizaron diversos productos cicatrizantes y antibacterianos previos a la aplicación de la Pasta de Unna Modificada, además de un método práctico de utilización de ésta. Del total de 23 pacientes estudiados, la úlcera más frecuente fue por insuficiencia venosa en 21 pacientes (91,30 por ciento). Se excluyeron 6 pacientes por presentar: vasculopatía periférica no venosa, reinfección, evolución tórpida del injerto o retiro de la consulta. Se incluyen 17 pacientes susceptibles de evaluar. La respuesta con cicatrización fue buena en 14 pacientes (82,35 por ciento). Nuestra experiencia demuestra que el reposo, limpieza y antibioticoterapia tópica y/o sistémica previa y el recambio semanal de la bota hasta lograr su epitelización o apta para colocar injerto dermoepidérmico son necesarios. Es un método efectivo para el tratamiento de las úlceras por insuficiencia venosa

Estudo da abrasividade de formulaçöes básicas de dentifrícios / Study of abrasivity of dentifrices basic formulations

O estudo da abrasividade de formulaçöes básicas de dentifrícios envolvendo vários tipos de espessantes e abrasivos, demonstrou uma dependência da interaçäo desses componentes, fundamentais nesse tipo de produto. As diferentes associaçöes estudadas tornaram possível a estruturaçäo de índices de abrasividade específicos a cada uma delas.

Viscosity of gums in vitro and their ability to reduce postprandial hyperglycemia in normal subjects

Experiments were carried out in vitro with three viscous polysaccharides (guar gum, pectin, and carboxymethylcellulose (CMC) of similar initial viscosity submitted to conditions that mimic events occurring in the stomach and duodenum, and their viscosity in these situations was compared to their actions on postprandial hyperglycemia in normal human subjects. Guar gum showed greater viscosity than the other gums during acidification and/or alkalinization and also showed larger effects on plasma glucose levels (35 per cent reduction in maximum rise in plasma glucose) and on the total area under the curve of plasma glucose (control: 20,314 + 1007 mg dl(-1) 180 min (-1) vs guar gum: 18,277 + 699 mg dl(-1) 180 min (-1), P<0.01). Pectin, which showed a marked reduction in viscosity at 37 degrees Celsius and after events mimicking those that occur in the stomach and duodenum, did not have a significant effect on postprandial hyperglycemia. The performance of viscosity and the glycemia response to CMC were at an intermediate level between guar gum and pectin. In conclusion, these data suggest that temperature, the process of acidification, alkalinization and exposure to intestinal ions induce different viscosity changes in gums having similar initial viscosity, establishing a direct relationship between a minor decrease of gum viscosity in vitro and a reduction of postprandial hyperglycemia.

Enhancement of liver size by stimulation of intact rat liver with exogenous hepatotrophic factors

Objetivos: Confirmar observaçoes anteriores (PARRA e col.) que sugeriam a possibilidade de fazer crescer o fígado em animais intactos, por estímulo hepatotrófico exógeno, ultrapassando seu tamanho biologicamente pré-determinado. Material e Métodos: Dois grupos de fêmeas de ratos Wistar, foram injetadas diariamente na cavidade peritoneal (40ml/kg) por sete dias consecutivos com as seguintes soluçoes: GRUPO A (controle) - soluçao salina com carboximetilcelulose (CMC) a 0,25 por cento; GRUPO B - soluçao de fatores hepatotróficos exógenos constituída de glicose, aminoácidos, insulina, glucagon, vitaminas, eletrólitos, triiodotironina e CMC na mesma concentraçao do grupo controle. No oitavo dia os animais forma sacrificados e avaliados os seguinte parâmetros: variaçao da massa hepática e DNA total do fígado dos animais do grupo experimental em relaçao ao grupo controle. Resultados: Foi observado crescimento da massa hepática de 114,16 +- 7,9 por cento acima do esperado e aumento do DNA total hepático de 12,99 +- 0,46 mg para 20,17 por fígado (p=0.0002). Conclusao: O aumento do tamanho do fígado a partir de sua massa primitiva, estimulada por fatores hepatotróficos exógenos, pode vir a ter aplicaçoes em intre-vivos, criando maior massa hepática a ser dividida entre doador e receptor.