Inhibition of astroglial hemichannels ameliorates infrasonic noise induced short-term learning and memory impairment.

As a kind of environmental noise, infrasonic noise has negative effects on various human organs. To date, research has shown that infrasound impairs cognitive function, especially the ability for learning and memory. Previously, we demonstrated that impaired learning and memory induced by infrasound was closely related with glia activation; however, the underlying mechanisms remain unclear. Connexin 43 hemichannels (Cx43 HCs), which are mainly expressed in hippocampal astrocytes, are activated under pathological conditions, lending support to the hypothesis that Cx43 HCs might function in the impaired learning and memory induced by infrasound. This study revealed that that blocking hippocampal Cx43 HCs or downregulating hippocampal Cx43 expression significantly alleviated impaired learning and memory induced by infrasound. We also observed that infrasound exposure led to the abundant release of glutamate and ATP through Cx43 HCs. In addition, the abundant release of glutamate and ATP depended on proinflammatory cytokines. Our finds suggested that the enhanced release of ATP and glutamate by astroglial Cx43 HCs may be involved in the learning and memory deficits caused by infrasound exposure.

The risk factors involved in airway mucus plug in children with ADV Pneumonia.

BACKGROUND: The risk factors for mucus plug in children with adenovirus (ADV) pneumonia. METHODS AND MATERIALS: A retrospective analysis was conducted of children diagnosed ADV pneumonia and underwent fiberoptic bronchoscopy admitted to the Xiamen Children's Hospital from September 2018 to September 2021.The patients were divided into a mucus plug group (39 cases) and a non-mucus plug group (53 cases). The children's data including sex, age, clinical presentation, laboratory test parameters, imaging and bronchoscopic data were collected. The risk factors for the development of airway mucus plug were analysed by multifactorial logistic regression. RESULTS: There were no statistically significant differences in sex, age, fever, hospitalization days, mixed infection, white blood cells (WBC) count, percentage of neutrophils (NE%), C-reactive protein(CRP), and D-dimer (all P > 0.05); Thermal range, procalcitonin (PCT), lactate dehydrogenase (LDH), Pleural effusion and associated decreased breath sounds was significantly higher in mucus plug group than in non-mucus plug group, and the differences were statistically significant (all P < 0.05); multifactorial logistic regression analysis showed that the duration of fever, PCT, and LDH were independent risk factors for the formation of mucus plugs. The critical values of ROC curves were pyroprocedure ≥ 6.5 d, PCT ≥ 0.705 ng/ml and LDH ≥ 478.5 U/L. CONCLUSION: Duration of fever, PCT and LDH levels were the independent risk factors for the formation of an airway mucus plug in children with ADV pneumonia.

Association between MTHFR c.677C>T variant and erectile dysfunction among males attending fertility clinic.

ABSTRACT: Genetic risk factors have been shown to contribute to the development of sexual dysfunction. However, the role of methylenetetrahydrofolate reductase (MTHFR) gene variants in the risk of erectile dysfunction (ED) remains unclear. In this study, we recruited 1254 participants who underwent ED assessed by the International Index of Erectile Function-5. The MTHFR c.677C>T variant was also measured by fluorescence polymerase chain reaction (PCR). No significant difference in the genotypic frequency of the MTHFR C677T polymorphism (CC, CT, and TT) was observed between men from the ED and non-ED groups. In addition, on binary logistic regression analysis, both crude and adjusted models showed that the risk of ED was not significantly associated with the C677T polymorphism. Interestingly, a significantly higher frequency of the 677TT polymorphism was found in severe and moderate ED (P = 0.02). The positive correlation between the MTHFR 677TT polymorphism and severe ED was confirmed by logistic regression analysis, even after adjusting for potential confounders (odds ratio [OR] = 2.46, 95% confidence interval [CI]: 1.15-5.50, P = 0.02). These findings suggest a positive correlation between the MTHFR 677TT polymorphism and the risk of severe ED. Identification of MTHFR gene polymorphisms may provide complementary information for ED patients during routine clinical diagnosis.

Time-trend analysis of tuberculosis diagnosis in Shenzhen, China between 2011 and 2020.

Objective: To describe the trend of tuberculosis (TB) diagnosis in the migrant city Shenzhen, China, and analyze the risk factors of diagnosis delays. Methods: Demographic and clinical information of TB patients from 2011 to 2020 in Shenzhen were extracted. A bundle of measures to enhance TB diagnosis had been implemented since late 2017. We calculated the proportions of patients who underwent a patient delay (>30 days from syndrome onset to first care-seeking) or a hospital delay (>4 days from first care-seeking to TB diagnosis). Multivariable logistic regression was used to analyze the risk factors of diagnosis delays. Results: During the study period, 43,846 patients with active pulmonary TB were diagnosed and registered in Shenzhen. On average, the bacteriological positivity rate of the patients was 54.9%, and this increased from 38.6% in 2017 to 74.2% in 2020. Overall, 30.3 and 31.1% of patients had a patient delay or a hospital delay, respectively. Molecular testing significantly increased bacteriological positivity and decreased the risk of hospital delay. People >35 years old, the unemployed, and residents had a higher risk of delays in both patient care-seeking and hospital diagnosis than younger people, workers, or migrants. Compared with passive case-finding, active case-finding significantly decreased the risk of patient delay by 5.47 (4.85-6.19) times. Conclusion: The bacteriological positivity rate of TB patients in Shenzhen increased significantly but the diagnosis delays were still serious, which may need more attention when active case-finding in risk populations and optimization of molecular testing.

Molecular Spectrum of α- and ß-Thalassemia among Young Individuals of Marriageable Age in Guangdong Province, China.

Thalassemia is a group of genetically heterogeneous diseases characterized by hemolytic anemia. To investigate molecular characteristics of α- and ß-thalassemia among young individuals of marriageable age in Guangdong Province, 24,788 subjects with suspected thalassemia were genetically tested for α- and ß-thalassemia by Gap-PCR and reverse dot blot during 2018-2019. For suspected rare thalassemia cases, DNA sequencing was performed to identify rare and unknown thalassemia gene mutations. A total of 14,346 thalassemia carriers were detected, including 7,556 cases of α-thalassemia with 25 genotypes and 8 α-gene mutations identified, 5,860 cases of ß-thalassemia with 18 genotypes and 18 ß-gene mutations identified, and 930 cases of compound α/ß-thalassemia. Among them, the frequency of -- SEA deletion was the highest in α-thalassemia (66.01%), followed by -α 3.7 (17.98%) and -α 4.2 (8.22%), and the frequency of CD41-42 (-TCTT) mutation was the highest in ß-thalassemia (38.38%), followed by IVS-II-654 (C > T) (25.67%), -28 (A > G) (15.76%), and CD17 (10.01%). In addition, 5 rare mutations (--THAI and HKαα, CD113, -90, and CD56) were found in the study population. Our results revealed molecular epidemiological background of α- and ß-thalassemia in Guangdong Province, which can support optimization of thalassemia prevention and control strategies. We demonstrated that thalassemia is heterogeneous with significant geographical differences and population specificity.

Effectiveness of Using Mean Corpuscular Volume and Mean Corpuscular Hemoglobin for Beta-thalassemia Carrier Screening in the Guangdong Population of China.

Beta (ß)-thalassemia is one of the most common hemoglobinopathies worldwide, creating major public health problems and social burdens in many regions. Screening for ß-thalassemia carriers is crucial for controlling this condition. To investigate the effectiveness of mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) for screening ß-thalassemia, retrospective data were analyzed for 6,779 ß-thalassemia carriers subjected to genetic testing following thalassemia screening in Guangdong province between January 2018 and December 2019. Prevalent mutations observed included CD41/42 (-TTCT) (38.43%), IVS-II-654 (C > T) (25.71%), -28 (A > G) (15.78%), CD17 (AAG > TAG) (10.03%), and ß E (GAG > AAG) (3.13%). In the ß 0, ß +, and HbE groups, MCV values were 63.8 ± 4.2 fL, 67.0 ± 5.5 fL, and 75.8 ± 5.6 fL, while MCH values were 20.1 ± 1.4 pg, 21.2 ± 1.9 pg, and 24.8 ± 2.0 pg, respectively. Among ß-thalassemia carriers, 85 (1.25%) and 28 (0.41%) individuals had MCV ≥ 80 fL and MCH ≥ 27 pg, respectively. Using a combination of MCV and MCH reduced the number of false negative screenings to 15 (0.22%). Therefore, evaluating both MCV and MCH is strongly recommended for screening ß-thalassemia carriers.

Quality of and Recommendations for Relevant Clinical Practice Guidelines for COVID-19 Management: A Systematic Review and Critical Appraisal.

Background: The morbidity and mortality of coronavirus disease 2019 (COVID-19) are still increasing. This study aimed to assess the quality of relevant COVID-19 clinical practice guidelines (CPGs) and to compare the similarities and differences between recommendations. Methods: A comprehensive search was conducted using electronic databases (PubMed, Embase, and Web of Science) and representative guidelines repositories from December 1, 2019, to August 11, 2020 (updated to April 5, 2021), to obtain eligible CPGs. The Appraisal of Guidelines for Research and Evaluation (AGREE II) tool was used to evaluate the quality of CPGs. Four authors extracted relevant information and completed data extraction forms. All data were analyzed using R version 3.6.0 software. Results: In total, 39 CPGs were identified and the quality was not encouragingly high. The median score (interquartile range, IQR) of every domain from AGREE II for evidence-based CPGs (EB-CPGs) versus (vs.) consensus-based CPG (CB-CPGs) was 81.94% (75.00-84.72) vs. 58.33% (52.78-68.06) in scope and purpose, 59.72% (38.89-75.00) vs. 36.11% (33.33-36.11) in stakeholder involvement, 64.58% (32.29-71.88) vs. 22.92% (16.67-26.56) in rigor of development, 75.00% (52.78-86.81) vs. 52.78% (50.00-63.89) in clarity of presentation, 40.63% (22.40-62.50) vs. 20.83% (13.54-25.00) in applicability, and 58.33% (50.00-100.00) vs. 50.00% (50.00-77.08) in editorial independence, respectively. The methodological quality of EB-CPGs were significantly superior to the CB-CPGs in the majority of domains (P < 0.05). There was no agreement on diagnosis criteria of COVID-19. But a few guidelines show Remdesivir may be beneficial for the patients, hydroxychloroquine +/- azithromycin may not, and there were more consistent suggestions regarding discharge management. For instance, after discharge, isolation management and health status monitoring may be continued. Conclusions: In general, the methodological quality of EB-CPGs is greater than CB-CPGs. However, it is still required to be further improved. Besides, the consistency of COVID-19 recommendations on topics such as diagnosis criteria is different. Of them, hydroxychloroquine +/- azithromycin may be not beneficial to treat patients with COVID-19, but remdesivir may be a favorable risk-benefit in severe COVID-19 infection; isolation management and health status monitoring after discharge may be still necessary. Chemoprophylaxis, including SARS-CoV 2 vaccines and antiviral drugs of COVID-19, still require more trials to confirm this.

Plastic bronchitis associated with adenovirus serotype 7 in children.

BACKGROUND: Plastic bronchitis is an uncommon but severe respiratory disease characterized by formation of casts in tracheobronchial tree. It can lead to airway obstruction and even respiratory failure. CASE PRESENTATION: Plastic bronchitis is mostly seen in both post-cardiac surgery patients, especially Fontan procedure, and infections including those caused by influenza viruses, Mycoplasma pneumoniae or tuberculosis. But it has rarely been reported to be associated with adenovirus infection. We report 2 cases of plastic bronchitis arising from adenovirus serotype 7 infection, manifested in repeated high fever, cough, and progressive dyspnea, and were diagnosed and eventually cured by bronchoscopy. CONCLUSIONS: Plastic bronchitis is a rare, variable and potentially fatal disease. In the cases we described, the cause was associated with adenovirus serotype 7 and its treatment required intervention with bronchoscopy and adequate control of the underlying disease.

Differential gene expression profile analysis in corticosterone-treated PC12 cells.

Major depressive disorder (MDD) is a highly prevalent psychiatric disorder that has been ranked as the 4th leading cause of disability worldwide. Past clinical and laboratory evidence has confirmed that abnormalities of the hypothalamic-pituitary-adrenal (HPA)-axis are involved in MDD development. In this study, we took advantage of corticosterone treatment of PC12 cells as a model to identify genes regulated by HPA-axis hormones. Next-generation RNA-Seq technology was utilized to explore genome-wide differentially expressed gene profiles between control and corticosterone treated PC12 cells. 1,274 genes with at least two-fold expression level change were identified. Gene ontology analysis showed that the top enriched biological processes included response to glucocorticoid signaling, apoptosis, cell division/DNA replication, and neuron projection/axon guidance, highly consistent with phenotypes of PC12 cells treated with corticosterone. Taken together, RNA-seq data is reliable and comprehensive, thus providing a valuable resource for understanding underlying mechanisms of glucocorticoid-induced neuron malfunction.

[Rapid Determination of Dodecylmorpholine in Aqueous Solutions with UV-Vis Spectrophotometry].

A rapid UV-Vis spectrophotometric method was proposed to determine the concentration of DMP in aqueous solutions. The linear concentration range of DMP solution at the range of 250～400 nm is 0.5～70 mmol·L(-1). At 275 nm, the linear fitting equation is A=0.030 7c+0.133 0 with a correlation of 0.980 9. The detection limitation is 9.46×10-5 mmol·L-1, the RSD (n=6) of the method were at the range of 0.100%～0.612%. The recovery ratio for salt solutions sample is 95%～104%. Temperature, pH, and coexisting K(+), Na(+), Mg(2+), Cl(-), Br(-), I(-), SO(2-)(4) ions do not affect the detection. The coexisting CO(2-)(3) and HCO(-)(3) ions can be eliminated with acidification. The results showed that the proposed method is simple in pretreatment process and has high accuracy and precision. It is a quick measurement method of DMP concentration in water solution, and can be used to measure DMP concentration in reverse flotation tail liquid and reverse flotation material pulp.

[Body mass, energy budget and leptin of mice under stochastic food restriction and refeeding].

Periods of restricted food intake that lead to lower body weight are often followed by rapid regaining of the lost weight after ad libitum refeeding, an event generally known as the "compensatory growth". To explore the physiological mechanisms underlying "compensatory growth", we evaluated a series of energetic parameters (energy intake, energy expenditure, body composition and serum leptin levels) of adult KM mice subjected to three cycles of stochastic food restriction following by ad libitum refeeding (SFR-Re). The results indicated that animals lost their body mass after stochastic food restriction and then regained to the control level after refeeding. After the final cycle, SFR-Re mice showed higher basal metabolic rate, lower nonshivering thermogenesis, and their cytochrome c oxydase activities, as well as uncoupling protein 1(UCP1) contents of brown adipose tissue all decreased compared with controls. Meanwhile, higher energy intake and digestibility, as well as heavier fat pads also were found in SFR-Re mice. But, serum leptin levels showed no difference between SFR-Re and control mice. On the whole, these findings indicated that when food is resourceful, SFR-Re mice are under rapid "compensatory growth" by increasing their food intake and energy storage efficiency, meanwhile, decreasing energy consumption in thermogenesis. Moreover, leptin may be a possible player in the regulations of energy budget and thermogenesis during "compensatory growth".

[Chromosome abnormalities and Y chromosome microdeletions in patients with the azoospermia and cryptozoospermia].

OBJECTIVE: To study the incidence of the chromosome abnormalities and Y chromosome microdeletions in Chinese patients with azoospermia and cryptozoospermia. METHODS: Conventional chromosomal karyotyping was used to analyze the chromosome abnormalities. Genomic DNA was extracted from peripheral blood samples and multiplex polymerase chain reactions (PCR) analyses were performed using specific primers to confirm the presence or absence of Y chromosome microdeletions. A total of 997 patients with azoospermia and cryptozoospermia were enrolled in the study. RESULTS: The incidence of chromosome abnormalities in the patient with azoospermia and cryptozoospermia was 28.4%. The major abnormal karyotypes included 47,XXY, 46,XY (Y>G), 46,XX, chimera and translocations. The incidence of the Y chromosome microdeletions was 17.4%. They were mainly found in the karyotypes of 46,XY and 46,XY (Y>G). CONCLUSION: Chromosome abnormalities were the most common hereditary causes of the patients with azoospermia and cryptozoospermia. The incidence of Y chromosome microdeletion was higher in the patients with karyotype of 46,XY and 46,XY (Y>G). Therefore, detection of the AZF microdeletion in these patients is helpful to determine the etiology and avoid the unnecessary treatment and vertical transmission of the genetic defects.

Attenuation of myocardial apoptosis by alpha-lipoic acid through suppression of mitochondrial oxidative stress to reduce diabetic cardiomyopathy.

BACKGROUND: Cardiac failure is a leading cause of the mortality of diabetic patients. In part this is due to a specific cardiomyopathy, referred to as diabetic cardiomyopathy. Oxidative stress is widely considered to be one of the major factors underlying the pathogenesis of the disease. This study aimed to test whether the antioxidant alpha-lipoic acid (alpha-LA) could attenuate mitochondrion-dependent myocardial apoptosis through suppression of mitochondrial oxidative stress to reduce diabetic cardiomyopathy. METHODS: A rat model of diabetes was induced by a single tail intravenous injection of streptozotocin (STZ) 45 mg/kg. Experimental animals were randomly assigned to 3 groups: normal control (NC), diabetes (DM) and DM treated with alpha-LA (alpha-LA). The latter group was administered with alpha-LA (100 mg/kg ip per day), the remainder received the same volume vehicle. At weeks 4, 8, and 12 after the onset of diabetes, cardiac apoptosis was examined by TUNEL assay. Cardiomyopathy was evaluated by assessment of cardiac structure and function. Oxidative damage was evaluated by the content of malondialdehyde (MDA), reduced glutathione (GSH) and the activity of manganese superoxide diamutase (Mn-SOD) in the myocardial mitochondria. Expression of caspase-9 and caspase-3 proteins was determined by immunohistochemistry and mitochondrial cytochrome c release was detected by Western blotting. RESULTS: At 4, 8, and 12 weeks after the onset of diabetes, significant reductions in TUNEL-positive cells, caspase-9,-3 expression, and mitochondrial cytochrome c release were observed in the alpha-LA group compared to the DM group. In the DM group, the content of MDA in the myocardial mitochondria was significantly increased, and there was a decrease in both the mitochondrial GSH content and the activities of Mn-SOD. They were significantly improved by alpha-LA treatment. HE staining displayed structural abnormalities in diabetic hearts, while alpha-LA reversed this structural derangement. The index of cardiac function (+/-dp/dtmax) in the diabetes group was aggravated progressively from 4 weeks to 12 weeks, but alpha-LA delayed deterioration of cardiac function (P < 0.05). CONCLUSIONS: Our findings indicate that the antioxidant alpha-LA can effectively attenuate mitochondria-dependent cardiac apoptosis and exert a protective role against the development of diabetic cardiomyopathy. The ability of alpha-LA to suppress mitochondrial oxidative damage is concomitant with an enhancement of Mn-SOD activity and an increase in the GSH content of myocardial mitochondria.

Mitochondrial gene mutations and type 2 diabetes in Chinese families.

BACKGROUND: Numerous mitochondrial DNA mutations are significantly correlated with development of diabetes. This study investigated mitochondrial gene, point mutations in patients with type 2 diabetes and their families. METHODS: Unrelated patients with type 2 diabetes (n = 826) were randomly recruited; unrelated and nondiabetic subjects (n = 637) served as controls. The clinical and biochemical data of the participants were collected. Total genome was extracted from peripheral leucocytes. Polymerase chain reaction, restriction fragment length polymorphism (PCR-RFLP) and cloning techniques were used to screen mitochondrial genes including np3316, np3394 and np3426 in the ND1 region and np3243 in the tRNA(Leu(UUR)). RESULTS: In 39 diabetics with one or more mitochondrial gene point mutations, the prevalence (4.7%, 39/826) of mtDNA mutations was higher than that (0.7%, 5/637) in the controls. The identical mutation was found in 23 of 43 tested members from three pedigrees. Affected family members presented with variable clinical features ranging from normal glucose tolerance to impaired glucose tolerance (IGT) (n = 2), impaired fasting glucose (IFG) (n = 1) to type 2 diabetes (n = 13) with 3 family members suffering from hearing loss. CONCLUSIONS: Type 2 diabetes in China is associated with several mitochondrial gene mutations. Aged patients with diabetic family history had a higher prevalence of mutation and various clinical pictures. Mitochondrial gene mutation might be one of the genetic factors contributing to diabetic familial clustering.

[High D-glucose alters PI3K and Akt signaling and leads to endothelial cell migration, proliferation and angiogenesis dysfunction].

OBJECTIVE: To investigate the effects of high D-glucose on migration, proliferation, and angiogenesis of endothelial cells and to make sure whether PI3K and Akt signaling pathway plays an important role in the pathogenesis of diabetic vascular complications. METHODS: Human umbilical vein endothelial cells (HUVECs) were cultured. D-glucose of the concentrations of 5 mmol/L, 15 mmol/L, and 30 mmol/L and mannitol of the concentrations of 5 mmol/L, 15 mmol/L, and 30 mmol/L were added in to the medium, the migration rate of the cells was measured by wound healing test and the cell proliferation was examined with CellTiter 96 AQ(ueous) One Solution cell proliferation assay. Matrigel was spread on 96-well plate, and culture of HUVECs with D-glucose and mannitol of different concentrations were added. Microscopic photography was used to calculate the total area of vascular bed, average vessel length, vessel number, branch points, so as to observe the angiogenesis. Immuno-precipitation was used to detect the expression of p85/PI3K. Western blotting was used to detect the protein expression of p85/PI3K, p-PI3K, GSK3beta (downstream kinase of Akt), p-Akt (Threonine308) and p-GSK3beta. LY294002, a PI3K inhibitor, of the concentrations of 0.1 micromol/L, 1 micromol/L, and 10 micromol/L was added into the culture fluid with 5 mmol/L D-glucose, then the endothelial cell migration, proliferation number, total area of blood bed, etc were observed. RESULTS: The migration rate of the 5 mmol/L D-glucose group was 100 +/- 23/microm2, and D-glucose dose-dependently decreased the migration rate, e.g. the migration rates of the 15 mmol/L and 30 mmol/L D-glucose groups were 77 +/- 18/microm2 and 46 +/- 18/microm2 respectively, both significantly lower than that of the 5 mmol/L D-glucose group (both P < 0.01). LY294002 of the concentrations of 0.1 micromol/L, 1 micromol/L, and 10 micromol/L dose-dependently decrease the endothelial cells migration rates to 68 +/- 16/microm2, 36 +/- 12/microm2, and 13 +/- 3/microm2 respectively (all P < 0.01) The cell proliferation rate of the 5 mmol/L, 15 mmol/L) and 30 mmol/L D-glucose groups were 59,128 +/- 7415/well, 33,144 +/- 9082/well, and 11,625 +/- 4196/well respectively, showing that D-glucose dose-dependently decreased the cell proliferation (all P < 0.01). LY294002 of the concentrations of 0.1 micromol/L, 1 micromol/L, and 10 micromol/L dose-dependently decrease the endothelial cell proliferation to 42,560 +/- 4213/well, 17,688 +/- 7198/well, and 5704 +/- 558/well respectively (all P < 0.01). 15 mmol/L and 30 mmol/L D-glucose decreased the numbers of total area of vascular bed, average tubule length, number of capillaries, and number of vessel branch pint formed on the Matrigel. LY294002 dose-dependently inhibited the angiogenesis (P < 0.05 or P < 0.01) 15 mmol/L and 30 mmol/L D-glucose dose-dependently inhibited the phosphorylation of p85/P13K and Akt (P < 0.05 and P < 0.01). However, D-glucose did not influence the protein expression of p85/PI3K and Akt. Mannitol did not influence the cell proliferation, angiogenesis, and the expression of p85/PI3K, phosphorylated p85/PI3K, Akt, phosphorylated Akt (Thr308), and phosphorylated GSK3beta. CONCLUSION: Hyperglycemia-impaired PI3K-Akt signaling may lead to migration, proliferation and angiogenesis dysfunction of endothelial cells in diabetes patients, which is likely to contribute to the pathogenesis of diabetic vascular complications.

[Study on the mitochondrial DNA mutations in patients with early-onset diabetes mellitus].

OBJECTIVE: To investigate the prevalence of mitochondrial DNA (mtDNA) mutations in patients with early-onset diabetes in Tianjin, and to explore the relationship between mtDNA mutations and diabetes. METHODS: 348 non-related patients whose age at onset of diabetes was less than 45 years were randomly recruited, and 207 non-related and non-diabetic subjects were enrolled as controls. All their clinical and biochemical data were collected. Total genome was extracted conventionally from the participants' peripheral leucocytes, and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and cloning techniques were applied to the screening of mtDNA mutations (including the 3316, 3394 and 3426 in ND1 region, 12026 in ND4 region, and tRNA [Leu(UUR)] 3243 A-->G mutation). RESULTS: The authors found 17 diabetics harboring the 12026 A-->G mutation in ND4 region (4.9%), 10 diabetics with mutations in ND1 region (including 5 diabetics with the 3394 T-->C mutation, 4 diabetics with 3316 G-->A mutation, one with 3426 A-->G mutation), and only two with the known 3243 A-->G mutation (0.6%). On the contrary, one control subject with the 3316 G-->A mutation, two with 3394 T-->C mutation and four with 12026 A-->G mutation were found. The prevalence of mtDNA mutations in the patient group is significantly higher than that in the control group (3.3%) (P<0.05). CONCLUSION: The above findings suggest that mtDNA mutation may be implicated in the pathogenesis of the examined diabetes.

HIV-1 Tat-mediated protein transduction of Cu,Zn-superoxide dismutase into pancreatic beta cells in vitro and in vivo.

Reactive oxygen species (ROS) are considered an important mediator in pancreatic beta cell destruction, thereby triggering the development of insulin-dependent diabetes mellitus. In the present study, we investigated the HIV-1 Tat protein transduction domain-mediated transduction of Cu,Zn-superoxide dismutase (SOD), which supplies SOD activity exogenously in pancreatic beta cells under oxidative stress. Tat-SOD fusion protein was successfully delivered into insulin-producing RINm5F cells and rat islet cells. The intracellular dismutation activities of SOD were found to increase in line with the amount of protein delivered into the cells. ROS, nitric oxide-induced cell death, lipid peroxidation, and the DNA fragmentation of insulin-producing cells were found to be significantly reduced when the cells were pretreated with Tat-SOD. Next, we examined the in vivo transduction of Tat-SOD into streptozotocin-induced diabetic mice. A single intraperitoneal injection of Tat-SOD resulted in the delivery of this biologically active enzyme to the pancreas. Moreover, increased radical scavenging activity in the pancreas was induced by multiple injections of Tat-SOD, and this enhanced the tolerance of pancreatic beta cells to oxidative stress. These results suggest that the transduction of Tat-SOD offers a new strategy for protecting pancreatic beta cells from destruction by relieving oxidative stress in ROS-implicated diabetes.

Dexamethasone-induced differentiation of pancreatic AR42J cell involves p21(waf1/cip1) and MAP kinase pathway.

Dexamethasone converts pluripotent pancreatic AR42J cells into exocrine cells expressing digestive enzymes. In order to address molecular mechanism of this differentiation, we have investigated the role of mitogen-activated protein (MAP) kinase pathway and gene expressions of p21(waf1/cip1) and nuclear oncogenes (c-fos and c-myc) during AR42J cell differentiation. Dexamethasone markedly increased the intracellular and secreted amylase contents as well as its mRNA level. However, cell growth and DNA content were significantly decreased. With these phenotypic changes, AR42J cells induced transient mRNA expression of p21(waf1/cip1) gene, which reached maximal level by 6 h and then declined gradually toward basal state. In contrast to p21(waf1/cip1), c-fos gene expression was transiently inhibited by 6 h and then recovered to basal level by 24 h. Increased c-myc expression detected after 3 h, peaked by 12 h, and remained elevated during the rest of observation. Dexamethasone inhibited epidermal growth factor-induced phosphorylation of extracellular signal regulated kinase. Inhibition of MAP kinase pathway by PD98059 resulted in further elevation of the dexamethasone-induced amylase mRNA and p21(waf1/cip1) gene expression. These results suggest that p21(waf1/cip1) and nuclear oncogenes are involved in dexamethasone-induced differentiation and inhibition of MAP kinase pathway accelerates the conversion of undifferentiated AR42J cells into amylase-secreting exocrine cells.