Validating Baboon Ex Vivo and In Vivo Radiation-Related Gene Expression with Corresponding Human Data.

The research for high-throughput diagnostic tests for victims of radio/nuclear incidents remains ongoing. In this context, we have previously identified candidate genes that predict risk of late-occurring hematologic acute radiation syndrome (HARS) in a baboon model. The goal of the current study was to validate these genes after radiation exposure in humans. We also examined ex vivo relative to in vivo measurements in both species and describe dose-response relationships. Eighteen baboons were irradiated in vivo to simulate different patterns of partial- or total-body irradiation (TBI), corresponding to an equivalent dose of 2.5 or 5 Sv. Human in vivo blood samples were obtained from patients exposed to different dose ranges: diagnostic computerized tomography (CT; 0.004-0.018 Sv); radiotherapy for prostate cancer (0.25-0.3 Sv); and TBI of leukemia patients (2 × 1.5 or 2 × 2 Sv, five patients each). Peripheral whole blood of another five baboons and human samples from five healthy donors were cultivated ex vivo and irradiated with 0-4 Sv. RNA was isolated pairwise before and 24 h after irradiation and converted into cDNA. Gene expression of six promising candidate genes found previously by us in a baboon model ( WNT3, POU2AF1, CCR7, ARG2, CD177, WLS), as well as three genes commonly used in ex vivo whole blood experiments ( FDXR, PCNA, DDB2) was measured using qRT-PCR. We confirmed the six baboon candidate genes in leukemia patients. However, expression for the candidate gene FDXR showed an inverse relationship, as it was downregulated in baboons and upregulated in human samples. Comparisons among the in vivo and ex vivo experiments revealed the same pattern in both species and indicated peripheral blood cells to represent the radiation-responsive targets causing WNT3 and POU2AF1 gene expression changes. CCR7, ARG2, CD177 and WLS appeared to be altered due to radiation-responsive targets other than the whole blood cells. Linear dose-response relationships of FDXR, WNT3 and POU2AF1 using human ex vivo samples corresponded with human in vivo samples, suggesting that ex vivo models for in vivo dose estimates can be used over a wide dose range (0.001-5 Sv for POU2AF1). In summary, we validated six baboon candidate genes in humans, but the FDXR measurements underscored the importance of independent assessments even when candidates from animal models have striking gene sequence homology to humans. Since whole blood cells represented the same radiation-responsive targets for FDXR, WNT3 and POU2AF1 gene expression changes, ex vivo cell culture models can be utilized for in vivo dose estimates over a dose range covering up to 3.5 log scales. These findings might be a step forward in the development of a gene expression-based high-throughput diagnostic test for populations involved in large-scale radio/nuclear incidents.

Lipogenesis in arterial wall and vascular smooth muscle cells of Psammomys obesus: its regulation and abnormalities in diabetes.

AIM: Lipogenesis is expressed in vascular smooth muscle cells (VSMCs), and such in situ lipogenesis could be providing the fatty acids for triglyceride synthesis and cholesterol esterification, and contributing to lipid accumulation in the arterial wall. This study investigated both the expression and regulation of lipogenesis in VSMCs to determine if they are modified in Psammomys obesus gerbils fed a high-fat diet as a model of insulin resistance and diabetes. METHODS: Aortas were collected from diabetic and non-diabetic P. obesus for histological examination, measurement of lipogenic gene expression and VSMC culture. RESULTS: The aortas of diabetic animals exhibited lipid deposits and foam cells as well as disorganization of elastic fibres. However, lipogenic gene expression was not modified. VSMCs in vitro from the aortas of diabetic animals had, compared with cells from non-diabetic animals, lower mRNA levels of SREBP-1c and ChREBP. An adipogenic medium stimulated moderate FAS and ACC1 expression in cells from both diabetic and non-diabetic animals, but glucose and insulin on their own had no such stimulatory action. Also, triiodothyronine (T3) had a clear stimulatory action, while angiotensin II had a moderate effect, in cells from non-diabetic P. obesus, but not from diabetic animals, whereas LXR agonists stimulated lipogenesis in cells from both animal groups. CONCLUSION: Lipogenesis is expressed in the arterial walls and VSMCs of P. obesus. However, its expression was not increased in diabetes, and did not respond to either T3 or angiotensin II. Therefore, lipogenesis in situ is unlikely to contribute to the accumulation of lipids in the arterial walls of diabetic P. obesus gerbils.

Lipase maturation factor 1: its expression in Zucker diabetic rats, and effects of metformin and fenofibrate.

AIM: High triglyceride (TG) levels are a risk factor for cardiovascular diseases, and TG concentrations depend on the balance between its appearance in and clearance from plasma. TG clearance is controlled mainly by lipoprotein lipase (LPL), the maturation and activity of which are dependent on lipase maturation factor 1 (LMF1) protein. The present study aimed to investigate LMF1 expression in hypertriglyceridaemia and the regulation of its expression, as little is currently known of these processes. METHODS: We measured LMF1 expression (mRNA) in Zucker diabetic rats (ZDF) throughout the development of obesity, insulin resistance and diabetes, and compared it with that of control rats. We also determined whether fenofibrate and metformin, agents with TG-lowering activities, have an effect on LMF1 expression in ZDF rats. RESULTS: At 7 weeks, the ZDF rats were obese, insulin-resistant and hypertriglyceridaemic, and their LMF1 mRNA levels were - whichever tissue was investigated - comparable to those of the control rats. Diabetic ZDF rats (14 and 21-week-old) also had high TG levels, but the presence of diabetes had no effect on LMF1 expression; mRNA levels remained comparable to those in the controls. Although fenofibrate and metformin both decreased plasma TG levels, fenofibrate had no effect on LMF1 expression, whereas metformin increased LMF1 mRNA in heart tissue (14- and 21-week-old ZDF rats; P<0.01), and induced a trend towards increases in adipose tissue (14-week-old ZDF rats) and muscle (14- and 21-week-old ZDF rats). CONCLUSION: LMF1 expression was not altered in this experimental animal model of obesity, insulin resistance and diabetes in the presence of raised TG levels. However, metformin increased LMF1 expression in the heart, suggesting that stimulation of LMF1 may play a part in its TG-lowering action.