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Long-term molecular and functional programming of adipose tissue by pre- and early postnatal malnutrition in a precocial sheep model

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  • Sharmila Ahmad
Over the past few decades, adipose tissue has become a focus of interest due to it involvements in various pathological diseases such as obesity and other metabolic disarrangements. Known as a tissue with higher degree of plasticity than other organs, it is very sensitive toward environmental cues, especially nutrition. Therefore, this study aimed to investigate long-term effects of pre-and postnatal malnutrition on the expandability and functional traits of four main adipose depots, namely subcutaneous (SUB), mesenteric (MES), perirenal (PER), and epicardial (EPI) adipose tissues at adolescence (6 months of age) and adulthood (2 ½ years old). To unravel the molecular and biological pathways underlying impaired adipose functionality and predisposition for metabolic diseases, we performed the Differential Expressed Gene (DEG) analysis of two of these adipose tissues (SUB and PER) and performed functional enrichment analysis using freely available software and established bioinformatics pipelines. In this study, adolescent lambs and adult sheep were born from dams (twin-pregnant) subjected to either normal (100% of energy and protein requirements; NORM), undernutrition (50% of energy and protein requirements; LOW) or overnutrition (150%/110% of energy/protein requirements; HIGH) in the last 6 weeks pre-partum (term~147 days). From day three post-partum, twin lamb were assigned to either a low-fat hay-based diet (CONV) or a high-carbohydrate-high-fat diet (HCHF) until 6 months of age, where subgroups of animals were then euthanized. The remaining animals were fed the same moderate low-fat diet for 2 years until adulthood, and were then euthanized at 2 ½ years of age. SUB was sampled above the longissimus dorsi muscle at the level of the 13th rib and EPI from the anterior surface of the heart, whereas MES and PER were randomly sampled after separation of the adipose tissue from their respective organs. Assessments of adipose tissue cellularity (i.e. proportion of adipocytes and membrane area) were performed using Visiopharm® software (Visiopharm®, Hoersholm, Denmark), mRNA expression of adipose-related markers (i.e. adipogenic, lipid metabolisms, inflammatory) were characterized by quantitative real-time polymerase chain reaction, and analyses for DEG (SUB and PER) applying deep RNA sequencing and it respective bioinformatic pipelines. By the end of adolescence (6-months old lambs), both prenatal LOW and HIGH predisposed for development of abdominal adiposity and extreme perirenal adipocyte hypertrophy by depressing non-obese cellularity in SUB and MES and by impairing SUB, MES, and PER hyperplasic (healthy) as opposed to hypertrophic (unhealthy) expandability, when challenged with postnatal HCHF diet-induced obesity. In PER, prenatal LOW (but not HIGH) upregulated inflammatory markers. The early postnatal HCHF diet induced hypertrophic expansion of adipose mass, particularly in PER, and in concomitant with upregulated inflammatory markers in all depots studied. In adulthood, depot- and sex specific long-term effects of pre-and postnatal nutrition was found, with sex-specific upper-limits for fat cell numbers and sizes (males<females) observed in SUB regardless of the early pre-postnatal nutrition, whereas PER was main target of prenatal nutrition, with reduced hypertrophic ability in LOW, and particularly LOW males, compared to NORM/HIGH males and all females. On the contrary, HIGH males had higher hypertrophic ability of PER, and thus phenotypically resembled all females. However, expression of most of the studied adipose markers was highest in LOW and LOW males, and could not account for the reduced in hypertrophic expandability in these groups. The EPI was mainly a target of the early postnatal HCHF diet with larger adipocytes size and upregulated expression of adipogenic and lipogenic markers observed in adulthood even after 2 years of dietary correction. The differential expression analysis of SUB revealed no DEG in any of the group comparisons, which confirms the upper-limits for expandability in SUB of adult sheep irrespective of the early nutrition history. However, in PER, 993 and 186 DEGs were identified between LOW versus HIGH and NORM, respectively, and these genes were enriched in biological pathways related to especially transmembrane transporter activity, motor activity related to cytoskeletal and spermatozoa function, cell cycle progression and response to extra cellular stimuli. The combination of pre-and postnatal malnutrition (LOW-HCHF and HIGH-HCHF) downregulated genes that participated in adipose remodeling and immunity-related pathways. The kidney cell differentiation was targeted by long-term effect of postnatal diet (HCHF versus CONV). In total, 6 and 2 hub genes were identified for prenatal LOW versus HIGH and NORM, respectively, whereas 8 and 9 hubs genes were detected for LOW-HCHF and HIGH-HCHF versus NORM-CONV. MMP9 was the only hubs gene for the postnatal diet (HCHF versus CONV). AURKA, MELK, TTK are among well-known markers to be expressed in adipose- and adipose stem cells, whose expression levels were altered by prolonged effects of particularly, prenatal LOW, whereas CTSS and IGTB2 are among adipose markers to be affected by LOW/HIGH-HCHF (versus NORM-CONV), and known to be linked to obesity. Our study suggests that there is a quite fixed upper-limit for SUB expandability in adulthood, which means that PER expandability may be a determining factor for early nutrition impacts on MES and perhaps ectopic (non-adipose tissue) fat deposition, rendering LOW male in particular predisposed for obesity-related metabolic risks. The EPI, however, was a particular target of the early postnatal HCHF diet with upregulated expression of adipogenic and lipogenic markers associated with adipocyte hypertrophy in adulthood. Both pre- and/or postnatal malnutrition differentially programmed (patho-) physiological pathways with implications for adipose functional development associated with metabolic dysfunctions, and PER was a major target.
Original languageEnglish
PublisherAarhus Universitet
Number of pages211
Publication statusPublished - Jan 2022

Note re. dissertation

Forsvar, onsdag den 5. januar 2022<br/>Termination date: 10.01.2022

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