Long-term infection passaging of Human Adenovirus 36 in monkey kidney cells

ABSTRACT Human Adenovirus 36 (HAdV-36) has been related to diverse effects on metabolism and may attenuate the lipid accumulation in kidneys with increased adiposity. Some of these effects would be related to viral persistence. However, until now, a model of persistent in vitro infection by HAdV-36 is unknown. In this study, we examined the cells of the Vero lineage to explore their permissiveness to long-term HAdV-36 infection. HAdV-36 was productively replicated in Vero cells and maintained long-term infection for up to 35 cell passages. A subculture was obtained from the cells that survived the primary infection at a low MOI (0.5). The production of the extracellular infectious virus with titers ranging from 104 to 106 TCID50/mL and DNA-bearing cells was detected. In long-term infected cells, the intracellular distribution of viral antigen was demonstrated by performing immunolocalization (IFI) and expression of cell-viral antigen in 50% of cells by flow cytometry, using anti-HAdV-36 hyperimmune rabbit serum. Furthermore, E1a and E4orf1 genes in long-term infected passages showed a decreasing trend. Our preliminary results reveal that renal epithelial monkey cells are permissive for the productive infection of HAdV-36. Vero cell culture long-term infection might be a promising model for addressing the fundamental aspects of the HAdV-36 biology that cannot reveal broadly-used cultures, which do not maintain long-term infection in primary or transformed cells.

Adenovirus 36 improves glycemic control and markers of Alzheimer's disease pathogenesis.

Alzheimer's disease (AD) is the most prevalent neurodegenerative disorder worldwide. While the causes of AD are unclear, several risk factors have been identified, including impaired glycemic control, which significantly increases the risk of cognitive decline and AD. In vitro and in vivo studies show that human adenovirus 36 (Ad36) improves glycemic control by increasing cellular glucose uptake in cells, experimental animal models and in humans who are naturally exposed to the virus. This study, tested improvement in glycemic control by Ad36 and delay in onset of cognitive decline in APPswe transgenic mice (Tg2576 line), a model of genetic predisposition to impaired glycemic control and AD. Three-month old APPswe mice were divided into Ad36 infected (Ad36) or mock infected (control) groups and baseline glycemic control measured by glucose tolerance test (GTT) prior to infection. Changes in glycemic control were determined 10- and 24-week post infection. Serum insulin was also measured during GTT. Cognition was determined by Y-maze test, while motor coordination and skill acquisition by rotarod test. Glycemic control as determined by GTT showed less deterioration in Ad36 infected mice over time, accompanied by a significant attenuation of cognitive decline. Analysis of brain tissue lysate showed significantly reduced levels of amyloid beta 42 in Ad36 mice relative to control mice. Golgi-Cox staining analysis also revealed reduced dendritic spines and synaptic gene expression in control mice compared to Ad36 infected mice. This proof of concept study shows that in a mouse model of AD, Ad36 improves glycemic control and ameliorates cognitive decline.

Potential role of E4orf1 protein in aging-associated impairment in glycemic control.

Aging constitutes a major risk factor for the development of type-2 diabetes (T2D) where glucose tolerance declines with age, resulting in a high prevalence of T2D and impaired glucose tolerance in the elderly population. Currently more than half of the 20 million U.S. adults with T2D are above the age of 60, and the largest increase in T2D prevalence is expected in the elderly. Obesity is a causative factor for T2D associated insulin resistance and hyperglycemia. Furthermore, the aging process is accelerated by hyperglycemia and effective treatment options are limited for the vulnerable aging population. One of the mechanisms contributing to aging associated hyperglycemia is resistance to insulin-mediated glucose disposal. Chronic hyperglycemia also accelerates aging by increasing pro-inflammatory milieu leading to impaired immune function. Although currently available anti-diabetic agents improve glycemic control, they have potential serious side effects in some cases. Therefore, additional and better drugs are urgently needed for treatment of insulin resistance and aging associated health risk factors. This review presents the novel use of a microbial protein, E4orf1 as a potential anti-diabetic agent, which functions independent of insulin and obesity, highlighting the role of unique sources for future drug development.

Twenty-five years of research about adipogenic adenoviruses: A systematic review.

Infectious etiology is implicated in chronic diseases such as gastric ulcer or atherosclerosis. However, "infection" is a recent term in the field of obesity. Since the first report in 1982 of obesity due to infection, several microbes have been linked to obesity. Among the adipogenic microbes, avian adenovirus SMAM-1 and human adenovirus Ad36 have been studied most extensively for the past 25 years. Here, we present a systematic review of literature about SMAM-1 and Ad36. Reports from North America, Europe, and Asia reveal strong evidence that Ad36 causes obesity in animals and paradoxically improves glycemic control, and in vitro data provides mechanistic explanation. Considering that experimental Ad36 infection of humans is unlikely, its causative role in human obesity or glycemic control has not been demonstrated unequivocally. Nonetheless, most, but not all, observational studies in children and adults link Ad36 infection to obesity and improvement in glycemic control. The E4orf1 gene of Ad36 was identified as responsible for better glycemic control. Overall, 25 years have considerably advanced knowledge about the role of infection in obesity. Potential translational benefits include the development of vaccines to prevent Ad36-induced obesity and drug development based on the E4orf1 protein to improve glycemic control.

High adenovirus 36 seroprevalence among a population of Hispanic American youth.

Infection with adenovirus 36 (Ad36) has been associated with risk of obesity in youth in some studies, but the seroprevalence of this virus has not been examined among all populations. As Hispanic-American youth are of greater risk for obesity than other American youth, we sought to determine the proportion of Ad36 seropositive (Ad36+) students in an urban middle school serving a Hispanic population. We further examined if Ad36+ students were more likely to have obesity, and if Ad36 serostatus impacted changes in weight status following a health intervention. We determined body mass index (BMI) at the beginning and end of a 16-week health intervention among 40 Hispanic-American middle-school students. Ad36 serostatus was determined by enzyme-linked immunsorbent assay (ELISA). Seventy percent of the students were Ad36+. Ad36+ and Ad36 seronegative (Ad36-) did not differ before or after the intervention in body weight measures. The odds of being classified as obese was 1.4 times greater among Ad36+ than Ad36- at baseline, and 2.4 times greater post-intervention, but these were not statistically significant. We report a high seroprevalence of Ad36 among a population of Hispanic-American students. Ad36 seropositivity was associated with a trend for a greater likelihood of having obesity, but did not impact response to a health intervention.

Role of LncRNA ROR in Ad36-induced browning of human adipose-derived stem cells / 中华内分泌代谢杂志

Objective To investigate the role of LncRNA ROR in Ad36-induced browning of human adipose-derived stem cells(hADSC). Methods After hADSC was induced by cocktail and Ad36 for 2,4,6,and 8 days,Oil red O staining was performed for observing the adipogenic status. The mRNA expressions of LncRNA ROR, uncoupling protein 1(UCP1),and PRDM16 were detected by real-time PCR and the protein expressions of UCP1 and PRDM16 were detected by Western-blot. After LncRNA ROR was knocked down by siRNA, UCP1 and PRDM16 mRNA and protein expression levels in the process of Ad36-induced adipocyte differentiation were detected by real-time PCR and Western-blot. Results Oil red O staining showed that fat droplets in the cocktail-induced group were larger than those in the Ad36-induced group. Compared with the cocktail group,the mRNA expressions of LncRNA ROR, UCP1 and PRDM16, and the protein expressions of UCP1 and PRDM16 in Ad36 group were significantly increased(P<0.05). The mRNA and protein expressions of UCP1 and PRDM16 in LncRNA ROR knockdown group were significantly lower than those in control group(P<0.05). Conclusion In the process of Ad36-induced hADSC differentiation,the up-regulation of LncRNA ROR may stimulate UCP1 and PRDM16 expression,and thus promote the browning of hADSC.

Regulation of PPARγ and CIDEC expression by adenovirus 36 in adipocyte differentiation.

This study is to investigate the role of adenovirus 36 (Ad36) in regulating expression of peroxisome proliferator-activated receptor γ (PPARγ) and cell death-inducing DFFA-like effector c (CIDEC) in Ad36-induced adipocyte differentiation. Human adipose-derived mesenchymal stem cells (hAMSCs) were isolated and cultured, and then infected with Ad36. Ad36-induced adipocytes were identified using quantitative real-time PCR and Oil red O staining. The expression levels of PPARγ and CIDEC in Ad36-induced adipocytes were determined by quantitative real-time PCR and Western blot analysis. Glucose uptake and intracellular triglyceride content were also determined in these induced cells. Our results from the Oil red O staining showed that Ad36 induced the differentiation of hAMSCs into human adipocytes in vitro. Moreover, the medium glucose concentration was significantly decreased, while the intracellular triglyceride content was significantly increased, in the Ad36-induced adipocytes, compared with the control group. Furthermore, our results showed that, the mRNA and protein expression levels of PPARγ and CIDEC were significantly upregulated in Ad36-induced adipocytes, in a time-dependent manner. On the other hand, compared with the control group, the CIDEC expression was downregulated when the Ad36-induced adipocytes were treated with the PPARγ inhibitor, GW9662. Ad36 could upregulate the expression level of CIDEC through increasing PPARγ expression during the adipocyte differentiation process.

The regulation of adenovirus type 36 infection and progranulin expression in Uygur obese patients / 西安交通大学学报(医学版)

ABSTRACT:Objective To investigate the regulation of adenovirus type 36 infection on precursor protein particles progranulin expression in Uygur obese patients.Methods Based on the diagnostic criteria of obesity,the samples were divided into obese group and non-obese group.Serum neutralization test was used to detect the antibody of Ad36.The progranulin mRNA expressions in abdominal omental and subcutaneous adipose tissues were detected by real-time quantitative PCR.ELISA method was used to determine serum progranulin protein levels. CD68 protein expression of macrophage was detected by immunohistochemistry. Results ① In the Uygur population of our study,compared with that in the non-obese group (38/1 1 7,32.5%),the number of obese patients (54/1 1 5,47.0%)infected with Ad36 was significantly higher than that in non-obese group (P <0.05 ).② Serum progranulin was significantly increased in the Ad36-infected obese group (408.45±1 56.92)than in non-obese group (326.1 1±1 58.60)(P <0.05).The mRNA expression of progranulin did not differ between the two groups.③ The macrophage infiltration was significantly higher in the Ad36-infected obese group (14 730.1 6 ± 2 227.39 )than in non-obese group (10 786.50 ± 2 772.80 )(P < 0.05 ).Conclusion Ad36 infection may be associated with the occurrence of obesity in Uygur population,and adenovirus type 36 infection may regulate the expression of serum progranulin at the protein level.

Human adenovirus-36 is uncommon in type 2 diabetes and is associated with increased insulin sensitivity in adults in Sweden.

BACKGROUND: Human adenovirus-36 (Adv36) increases adiposity, but also upregulates distal insulin signaling in vitro in human adipose and muscle tissue and in vivo in the rodent independently of adiposity. Accordingly, healthy adults and children with antibodies against Adv36 had increased insulin sensitivity and reduced hepatic lipid accumulation. We hypothesized that Adv36 infection would be less frequent in individuals with type 2 diabetes or impaired glycemic control. METHODS: Presence of antibodies against Adv36 was analyzed for association to type 2 diabetes or impaired glycemic control in a two-wave population-based sample of well-characterized adults (n = 1734). Indices of impaired glycemic control included oral glucose tolerance, and circulating fasting levels of glucose, insulin, and insulin-like growth factor binding protein-1 (IGFBP-1). RESULTS: Adv36 seropositivity was more common in those with normal glucose tolerance (NGT) than in those with diabetes (females: OR 17.2, 95% CI 4.0-74.3; males: OR 3.5, 95% CI 1.8-6.7). Also, females with NGT had higher frequency of Adv36 seropositivity than females with prediabetes (impaired glucose tolerance and/or impaired fasting glucose; OR 1.8, 95% CI 1.1-3.1). Within the female prediabetes group Adv36 seropositivity was associated with higher insulin sensitivity reflected by reduced HOMA-IR and increased IGFBP-1. CONCLUSION: Adv36 infection is associated with lower occurrence of type 2 diabetes and better insulin sensitivity in adults, particularly among females.

Harnessing the beneficial properties of adipogenic microbes for improving human health.

Obesity is associated with numerous metabolic comorbidities. Weight loss is an effective measure for alleviating many of these metabolic abnormalities. However, considering the limited success of most medical weight-management approaches in producing a sustained weight loss, approaches that improve obesity-related metabolic abnormalities independent of weight loss would be extremely attractive and of practical benefit. Metabolically healthy obesity supports the notion that a better metabolic profile is possible despite obesity. Moreover, adequate expansion of adipose tissue appears to confer protection from obesity-induced metabolic comorbidities. To this end, the 10th Stock conference examined new approaches to improve metabolic comorbidities independent of weight loss. In particular, human adenovirus 36 (Ad36) and specific gut microbes were examined for their potential to influence lipid and glucose homeostasis in animals and humans. While these microbes possess some undesirable properties, research has identified attributes of adenovirus Ad36 and gut microbes that may be selectively harnessed to improve metabolic profile without the obligatory weight loss. Furthermore, identifying the host signalling pathways that these microbes recruit to improve the metabolic profile may offer new templates and targets, which may facilitate the development of novel treatment strategies for obesity-related metabolic conditions.