Browsing by Author "Ramalingam, Latha"
Now showing 1 - 14 of 14
- Results Per Page
- Sort Options
Item A theory-based nutrition education intervention to enhance complementary feeding practices among young children in Southern Ethiopia(2021-05) Moyeda Carabaza, Ana Florencia; Murimi, Mary; Dawson, John A.; Ramalingam, Latha; Thompson, Leslie; Vipham, JessiePurpose. The purpose of this study was two-fold: 1) to assess the nutritional status of children; their complementary feeding practices; household food insecurity; mothers’ knowledge and attitudes towards complementary feeding practices; and water, sanitation and hygiene practices; and 2) to design and implement a nutrition education intervention (NEI) to improve child feeding practices and related nutritional status by improving mothers’ knowledge, attitudes, and practices (KAP) of complementary feeding for children aged 6 to 23 months in the Sidama and Oromia regions of Ethiopia. Methods. This study used cluster-randomized trial design with an intervention and a control group. Pairs of mothers and their children aged six to 23 months from six villages located in Arsi Negele, Wondo Genet, and Dale districts were recruited and randomly assigned into the intervention (IG, n = 99) or control group (CG, n = 91). The NEI was was guided by the Social Cognitive Theory. The NEI curriculum spanned a period of six weeks, with a weekly three-hour session. Each session was designed to provide a lecture on the week’s topic, a discussion using counseling cards, a cooking demonstration, and key messages. The control group did not receive any intervention. Data was collected pre- and post-intervention using the knowledge, attitudes and practices towards complementary feeding questionnaire and anthropometric measurements. Household food insecurity; water, sanitation, and hygiene practices; and demographic characteristics were assessed only at baseline. Linear and generalized mixed models were used to assess the effectiveness of the NEI. Linear and logistic regressions were performed to assess associations between variables at baseline and post-intervention. Results: A quarter (25.8%) of children were stunted, 13.7% were underweight, and 6.8% were wasted. Most (87.4%) children achieved the recommended minimum meal frequency (MMF), while only 27.4% achieved the minimum dietary diversity (MDD) and 26.8% the minimum acceptable diet (MAD). Majority (80%) of households were food insecure, with 40% of households reporting severe food insecurity. At baseline, half (50%) of mothers demonstrated poor levels of knowledge on complementary feeding practices. Mothers that lacked formal education and that lived in moderate and severely food insecure households had a reduced probability of achieving children’s MDD (p < .05) and MAD (p < .05). Mothers with low-income levels were less likely to achieve children’s MAD (p = .014). At post-intervention, the IG significantly increased its knowledge scores compared to the CG (3.47 ± 1.28 vs. 0.57 ± 1.41, p = .004). Similarly, the IG had a significantly greater increase in the proportion of children that achieved the MDD (p = .003) and MAD (p < .001) than the control group. Post-intervention results demonstrated a significant reduction of stunting in the IG than those in the CG (p = .035). Conclusion: A high prevalence of child undernutrition, food insecurity, and suboptimal complementary feeding practices remains in the region. In addition, this study demonstrated that theory-based NEIs that use low-literacy strategies are effective in improving complementary feeding knowledge and practices, and the nutritional status of children. Future NEIs that focus on the improvement of complementary feeding practices in Ethiopia should implement low-literacy strategies such as the use of plain language, applying the teach-back method, integrating activities that are based in observational learning, and providing an intensive training for nutrition educators.Item C‐peptide as a therapy for type 1 diabetes mellitus(2021) Washburn, Rachel L. (TTUHSC); Mueller, Karl (TTUHSC); Kaur, Gurvinder (TTUHSC); Moreno, Tanir (TTUHSC); Moustaid‐moussa, Naima (TTU); Ramalingam, Latha; Dufour, Jannette M. (TTUHSC)Diabetes mellitus (DM) is a complex metabolic disease affecting one‐third of the United States population. It is characterized by hyperglycemia, where the hormone insulin is either not produced sufficiently or where there is a resistance to insulin. Patients with Type 1 DM (T1DM), in which the insulin‐producing beta cells are destroyed by autoimmune mechanisms, have a significantly increased risk of developing life‐threatening cardiovascular complications, even when exogenous insulin is administered. In fact, due to various factors such as limited blood glucose measurements and timing of insulin administration, only 37% of T1DM adults achieve normoglycemia. Furthermore, T1DM patients do not produce C‐peptide, a cleavage product from insulin processing. C‐ peptide has potential therapeutic effects in vitro and in vivo on many complications of T1DM, such as peripheral neuropathy, atherosclerosis, and inflammation. Thus, delivery of C‐peptide in conjunction with insulin through a pump, pancreatic islet transplantation, or genetically engineered Sertoli cells (an immune privileged cell type) may ameliorate many of the cardiovascular and vascular complications afflicting T1DM patients.Item Effect of a MetAP2 Inhibitor on Adipogenesis(2018-05) Siddik, Ad Abu Bakkar; Hegde, Vijay; Rahman, Shaikh; Ramalingam, LathaObesity is one of the key factors in health related issues of our times and prevalence of obesity has nearly tripled since 1975. Angiogenesis is the process of blood vessel formation from endothelial cells or pre-existing vessels and closely associated with obesity or adipose tissue formation. So controlling angiogenesis can be a modulating factor for obesity. Methionine Amino Peptidase 2 (MetAP2) a metalloproteinase, has been shown be a regulatory element for angiogenesis and a target molecule for anti-angiogenic compounds. In our study we have investigated the effect of a MetAP2 inhibitor compound BL6 on adipogenesis. Our central hypothesis is that MetAP2 inhibitor BL6 will block adipogenesis in murine 3T3-L1 cells by blocking angiogenesis. Angiogenesis is also associated with tube formation of Human Umbilical Vein Endothelial Cells (HUVEC). Therefore, we also hypothesized that compound BL6 will also inhibit tube formation in HUVECs. Murine 3T3-L1 cells were treated with MetAP2 inhibitor compound BL6 (20µM, 50 µM and100µM) during differentiation with MDI cocktail. Cells were differentiated for 8 days followed by staining with Oil Red O to determine lipid accumulation. Protein and RNA was also extracted from these cells. Adiponectin, PPARγ, C/EBPα, C/EBP Beta, SREBP1 and FAS expression was quantified by Western Blot and RT-PCR. During in vitro differentiation of mouse 3T3-L1 preadipocytes, administration of 50μM and 100µM of BL6 resulted in reduced lipid accumulation (P<0.05). 100µM BL6 also resulted in decreased Adiponectin, PPAR γ, SREBP1 and FAS level though not significant. Surprisingly, elevated level of C/EBPα was observed inspite block to PPAR γ. Furthermore, cells treated with BL6 during the differentiation period maintained normal metabolic health and improved glucose uptake despite suppression of adipogenesis. Apart from block to adipogenesis, tube formation reduction in HUVECs was observed when administered with 20μM and 50μM of BL6. Collectively this proof of concept study supports the development of a MetAP2 inhibitor, BL6 as a putative therapeutic agent.Item Effects of fatty acid supplementation on gene expression, lifespan, and biochemical changes in wild type and mutant C. elegans strains(2019-08) Bouyanfif, Amal; Hequet, Eric; Moustaid-Moussa, Naima; Ballou, Michael A.; Mendu, Venugopal; Ramalingam, Latha; Koboziev, IiuriiSupplementation of diet with omega-3 fatty acids has proven to be beneficial and directly associated with human health development. Polyunsaturated fatty acids (PUFAs) generate pro-resolving lipid mediators during the resolution phase of acute inflammation. In this dissertation, we were interested in studying the effects of fatty acid supplementation on gene expression, lifespan, and biochemical changes in wild-type and mutant C. elegans strains. The choice of C. elegans for this investigation is based on the fact that, unlike mammals, C. elegans does not require essential fatty acids in its diet because it is capable of synthesizing PUFAs such as arachidonic acid and eicosapentaenoic acid using saturated and monosaturated fatty acids from bacteria as precursors. In addition, C. elegans mutant strains such as tub-1 and fat-3 are also available. In contrast to wild-type, mutant strain fat-3 lacks 6 desaturase activity and fails to produce any of the common C20 PUFAs while in mutant strain tub-1 the functional loss of tubby ortholog called tub-1/F10B5.5 in C. elegans leads to accumulation of triglycerides, which are the major form of stored fat. Our findings show that C. elegans is suitable model to study the effects of PUFAs on aging, gene expression, fatty acid metabolism, and biochemical composition changes. Quantitative PCR did not show significant effects of EPA supplementation, probably because worms were not synchronized. When worms were synchronized using microfluidic device, long-term consumption of PUFAs (eicosapentaenoic acid) resulted in accelerated aging due to peroxidation of the unsaturation but statistically not significant. Our exploratory investigation using Fourier Transform Infrared microspectroscopy was revealed to be interesting. The results showed that not only biochemical change (in lipids and proteins) occur during C. elegans lifespan, but also as a result of supplementation of the growth media with saturated and unsaturated fatty acids.Item Eicosapentaenoic acid regulates brown adipose tissue metabolism in high-fat-fed mice and in clonal brown adipocytes(Elsevier, 2017-01) Pahlavani, Mandana; Razafimanjato, Fitia; Ramalingam, Latha; Kalupahana, Nishan S.; Moussa, Hanna; Scoggin, Shane; Moustaid-Moussa, NaimaBrown adipose tissue (BAT) plays a key role in energy expenditure through its specialized thermogenic function. Therefore, BAT activation may help prevent and/or treat obesity. Interestingly, subcutaneous white adipose tissue (WAT) also has the ability to differentiate into brown-like adipocytes and may potentially contribute to increased thermogenesis. We have previously reported that eicosapentaenoic acid (EPA) reduces high-fat (HF)-diet-induced obesity and insulin resistance in mice. Whether BAT mediates some of these beneficial effects of EPA has not been determined. We hypothesized that EPA activates BAT thermogenic program, contributing to its antiobesity effects. BAT and WAT were harvested from B6 male mice fed HF diets supplemented with or without EPA. HIB 1B clonal brown adipocytes treated with or without EPA were also used. Gene and protein expressions were measured in adipose tissues and H1B 1B cells by quantitative polymerase chain reaction and immunoblotting, respectively. Our results show that BAT from EPA-supplemented mice expressed significantly higher levels of thermogenic genes such as PRDM16 and PGC1α and higher levels of uncoupling protein 1 compared to HF-fed mice. By contrast, both WATs (subcutaneous and visceral) had undetectable levels of these markers with no up regulation by EPA. HIB 1B cells treated with EPA showed significantly higher mRNA expression of PGC1α and SIRT2. EPA treatment significantly increased maximum oxidative and peak glycolytic metabolism in H1B 1B cells. Our results demonstrate a novel and promising role for EPA in preventing obesity via activation of BAT, adding to its known beneficial anti-inflammatory effects.Item Estimation of absorbed dose to adipose tissue from full field digital mammogram using Monte-Carlo simulation(2018-12) Rashid, Al Maqsudur; Moussa, Hanna; Moussa, Hanna; Kumar, Golden; Parameswaran, Siva; Abidi, Noureddine; Ramalingam, LathaRecent study has found influential role of inflamed adipose (fat) tissue in breast cancer due to low dose radiation. Various inflammatory factors secreted from these irradiated adipose cells might stimulate surrounding tissues including dormant cancer cells. Previously, only glandular tissues were considered most radio-sensitive organ in breast hence no dosimetry studies exist on breast adipose tissues. The objective of this study was full field digital mammogram (FFDM) radiation dosimetry in adipose tissue region of the breast. The average energy deposition in the adipose tissue was simulated as a function of different target-filter materials, quality of x-ray beam (half value layer), percentage of adipose tissue, compressed breast thickness, peak tube voltages (kVp), and tube current-time products (mAs). These variables were taken from recommended imaging techniques published in the technique charts by the manufacturer corresponding to patient’s anatomy and actual mammogram experiment data of anonymous patient’s courtesy. The absorbed doses were simulated and estimated using Monte Carlo N-Particle (MCNP 6) transport codes developed by Los Alamos national laboratory. The obtained dose information’s could be helpful for breast cancer research and provide data necessary for epidemiological studies to improve various risk models such as those presented in BEIR (biological effects of ionizing radiation) or ICRU (International Commission on Radiological Protection) reports.Item FTIR microspectroscopy study of compositional changes in biological samples(2017-08) Parajuli, Prakash; Abidi, Noureddine; Hequet, Eric; Moussa, Hanna; Ramalingam, LathaFourier Transform Infrared (FTIR) microspectroscopy has emerged as a powerful technique for analyzing biological samples. Obtaining spatially resolved molecular information from the samples is one of the major advantages of this technique. Qualitative, as well as quantitative analysis of the sample, is possible with the use of this technique. It gives information regarding the intrinsic molecular chemistry of the sample. It could be applied for the analysis of wide range of samples. In this study, we used FTIR microspectroscopy to investigate the changes in the chemical composition of mouse adipose tissue upon oxygen plasma treatment and cellulose substrate upon plasma treatment and grafting of vinyl laurate. The first chapter describes the instrumentation and principle of operation of FTIR microspectroscopy and microwave plasma. Moreover, it provides a description of adipose tissue and cellulose. The chapter also covers reactive oxygen species (ROS), their functions and their role in oxidative stress. In the second chapter, the effect of ROS on lipids and proteins of mouse white adipose tissue was studied using FTIR microspectroscopy. Microwave plasma was used to induce oxidative damage to the biomolecules of white adipose tissue (WAT) of mouse through insitu production of ROS. The analysis of the IR spectra extracted from the infrared (IR) images of adipocyte and ECM of the mouse white adipose tissue showed a significant effect of ROS on lipids. Mainly, unsaturated lipids were found to be highly affected by ROS as a drastic decrease in the area of the band attributed to the olefinic (=CH-) vibration from lipid was observed. Similarly, changes associated with saturated lipids were also observed. Along with the decrease in the area of bands assigned to lipids, significant increase in the area of carbonyl (C=O) band was observed. However, amide bands from proteins did not change significantly, indicating that proteins are comparatively more resistant to ROS than lipids. Chemimaps and band ratio images were developed from the FTIR image recorded after each treatment. Chemimaps clearly showed decreasing concentrations of olefinic group and increasing concentration of the carbonyl group. Similarly, band ratio images showed decreasing intensity of olefinic/lipid ratio and increasing intensity of carbonyl/lipid ratio. In the third chapter, grafting of vinyl laurate monomer on the regenerated cellulose film and cotton fibers was studied using FTIR microspectroscopy. The grafting of monomer on the cellulose films and cotton fibers was initiated by means of microwave plasma treatment. The analysis of spectra extracted from the FTIR images of the monomer grafted samples showed the presence of additional peaks at 1735, 2925 and 2855 cm-1. Comparison of correlation coefficient between the spectra of the control and those of treated samples showed significantly higher correlation in treated samples compared to control samples. Chemical distribution maps showed non-uniform distribution of vinyl laurate on the cellulose film as indicated by a non-uniform distribution of carbonyl group on the FTIR image of the cellulose film. In the final chapter, a brief summary of three chapters is included. It also describes the importance of FTIR imaging and future directions of the projects described in chapters 2 and 3. Overall, the results showed that FTIR imaging is effective in analyzing animal tissue as well as plant materials. This shows a wide range of application of FTIR imaging and could be used as a rapid and sensitive technique to monitor subtle changes in biomolecules. The simplicity of the sample preparation, non-destructiveness, and ease in obtaining results has made it an excellent tool for the analysis of biological samples. Overall, the present study is further important as it sheds light on the compositional changes occurring in the macromolecular content of biological samples, which could be useful for further studies.Item Hepatoprotective mechanisms of eicosapentaenoic acid in high-fat fed mice and identification of biomarkers for Non-Alcoholic Fatty Liver Disease in humans(2018-12) Albracht, Kembra; Moustaid-Moussa, Naima; Ramalingam, Latha; Wang, Shu; Robert-McComb, Jacalyn; Kalupahana, Nishan; Rahman, ShaikhNon-alcoholic fatty liver disease (NAFLD) is a rising epidemic, in part due to its association with obesity and obesity-related metabolic abnormalities. However, the relationship between body weight, NAFLD, and insulin resistance is not well characterized in humans, especially in populations that develop metabolic complications at lower body mass index, such as South Asians. Therefore, research that can lead to effective diagnostic as well as prevention/ treatment strategies for NAFLD and related diseases is greatly needed. Several studies in animal and humans used dietary interventions, such as omega-3 (n-3) polyunsaturated fatty acids (PUFAs), to reduce NAFLD as well as inflammation-related metabolic disorders. While the anti-inflammatory and hypotriglyceridemic properties of n-3 PUFAs are well-known, mechanisms mediating their benefits in liver and in NAFLD are less understood. Previous studies in our lab have shown reduced hepatic triglyceride accumulation, despite similar body weights, between high-fat fed mice and those supplemented with eicosapentaenoic acid (EPA), a metabolite of n-3 PUFA. Hence, our objectives were: 1. Determine metabolic, inflammatory, and molecular changes in serum biomarkers in humans with NAFLD, and determine whether these are independent of body weight; and 2. Determine mechanisms by which n-3 PUFA reduces fatty liver in diet-induced obese mice, independent of adiposity. In our human study, we found central adiposity, subcutaneous and visceral adipocyte area, fasting blood glucose, and serum resistin to be the best indicators of NAFLD in South Asian women. Moreover, in liver tissues from mice and in HepG2 human hepatoma cells cultured in an inflammatory environment, we demonstrated that EPA ameliorated high-fat diet-induced NAFLD and improved energy metabolism. This was mediated by reduced de novo lipogenesis and triglyceride synthesis and by up-regulated lipid catabolism. Furthermore, gene and protein markers of inflammation were reduced with EPA. Interestingly, we found these processes to be mediated, in part, by EPA-regulated microRNA (miRNA) known to affect these hepatic processes. In conclusion, this work contributed significant knowledge towards understanding the link between NAFLD, obesity, insulin resistance and inflammation. Moreover, our animal studies provide novel science-based rationale for EPA supplementation in NAFLD patients, especially in South Asian populations with metabolic complications that are independent of body weight.Item Mechanisms linking the adipocyte renin angiotensin system to obesity, inflammation and endoplasmic reticulum stress(2020-05) Menikdiwela, Kalhara R.; Moustaid-Moussa, Naima; Ramalingam, Latha; Kalupahana, Nishan S.; Dufour, Jannette; Wang, ShuThe Renin Angiotensin System (RAS), the primary regulator of blood pressure, has been linked to several metabolic disorders including obesity. Its components are highly expressed in various tissues and organs such as adipose, liver, and kidney. During obesity, levels of angiotensin II (Ang II) and angiotensinogen (Agt), the key components of RAS system, are significantly elevated. Accumulated evidence over the past few decades demonstrated that adipose overexpression of RAS components disrupts metabolic homeostasis in the adipose tissue promoting chronic environment leading to metabolic disorders. Obesity and its co-morbidities are reaching epidemic portion, therefore, research that can unravel mechanistic involvement of RAS in obesity is urgently needed. Finding effective treatment strategies for RAS-induced obesity will help us get one step closer to addressing the global health concerns associated with RAS (obesity, diabetes and cardiovascular diseases) which is the primary focus of this dissertation study. Animal models with RAS overexpression or inactivation provide excellent tools to understand the involvement of RAS in obesity. Agt overexpression specifically in mice adipose tissue, showed elevated expression of pro-inflammatory cytokines with lower amounts of anti-inflammatory cytokines. Additionally, in vivo and in vitro studies conducted in cardiovascular system have demonstrated that RAS component Ang II activates several physiological processes including endoplasmic reticulum (ER) stress, oxidative stress, autophagy and inflammation. However, the role of RAS overactivation in adipose tissue during obesity and how these processes are regulated, remain to be elucidated. MicroRNAs (miRNA) are potential mediators as they are capable of regulating multiple genes post-transcriptionally. Previous research conducted in our lab has reported that overexpression of Agt in adipose tissue (Agt-Tg) induces obesity, inflammation and ER stress through yet unidentified mechanisms. Accordingly, we hypothesized that adipose overactivation of angiotensinogen leads to stimulation of inflammation, and ER stress which could contribute to obesity. We further hypothesize that RAS-induced metabolic alterations are mediated via miRNAs. The two main objectives of this research were: (1) To determine the effects of RAS in adipose inflammation, and ER stress. Wild type (Wt), Agt-Tg and Agt-knockout (KO) mice along with clonal mice and human adipocytes were used to test this objective. We identified that RAS overactivation significantly activates the expression of ER stress, inflammation and autophagic markers. We also found that RAS increases ER stress in part via Nuclear factor kappa B- Yin Yang 1 (NF-κB-YY1) axis through Angiotensin II receptor type 1 (AT1) receptor. Furthermore, studies performed with RAS inhibitors such as angiotensin converting enzyme (ACE) inhibitor, angiotensin receptor blockers and KO mice showed reduced levels of pro-inflammatory markers. Corroborating these results, we reported that AT1 blocker, telmisartan reduced ER stress and inflammation in cells further confirming potential RAS function via AT1 receptor. (2) To determine mechanisms by which RAS induces metabolic alterations in adipose tissue and identify miRNAs mediating effects of RAS during obesity. From our mechanistic studies related miRNAs, we showed that several miRNAs were affected by RAS overactivation. Out of 31 differentially expressed miRNAs, miR-690 was significantly overexpressed in Agt Tg mice compared to wild type mice. Interestingly, miR-690 appears to have a protective role during RAS induced obesity, targeting mitogen-activated protein kinase kinase 3 (MAP2K3). We showed that miR-690 ameliorates downstream inflammatory and ER stress markers associated with p38MAPKs pathway by inhibiting MAP2K3 expression. In conclusion, this study contributed significantly to a better understanding of how RAS overactivation disrupts metabolic homeostasis in the adipose tissue during obesity. Adipose RAS overexpression affects signaling pathways (inflammation, ER stress and autophagy) and miRNAs which regulate these molecular pathways. Additionally, this work provides science-based rationale for detrimental effects of RAS overexpression in the adipose tissue, suggesting novel molecular targets that could be used in the development of future therapeutic.Item Mechanisms of effects of tart cherry in obesity, inflammation and aging(2020-05) Jayarathne, Shasika; Moustaid-Moussa, Naima; Ramalingam, Latha; Oldewage-Theron, Wilna; Thompson, Leslie; Zabet, Masoud; Park, Oak-HeeObesity is a complex disease, which has become a major health problem worldwide. Obesity is associated with adipose tissue hypertrophy and hyperplasia, as well as pathologic endocrine alterations of adipose tissue including local and systemic low-grade inflammation. Aging has a significant influence on economy, health and demography with healthy aging being a global challenge. Aging and health span are determined by both environmental and genetic factors. It is evident that obesity reduces cellular and molecular functions and may lead to aging process as well. Additionally, pathological differences during obesity such as inflammation and oxidative stress as well as cellular damage may accelerate the rate of aging. Therefore, there is a close association between aging and obesity. The insulin/insulin-like growth factor-1 (IGF-1) signaling (IIS) pathway is a key mediator of aging in C. elegans and mammals. Specifically, DAF-2 signaling, an ortholog of human IGF, controls DAF-16/FOXO transcription factor, a master regulator of metabolism and longevity. Moreover, mitochondrial dysfunction and oxidative stress are both linked to aging. Dietary interventions are among effective non-genetic or pharmacological means to combat both obesity and aging. In this dissertation, we primarily focused on how nutritional interventions can be used to reduce adipose tissue inflammation associated with obesity and aging. Basic and clinical studies demonstrate that foods containing bioactive compounds are capable of preventing both obesity and adipose tissue inflammation, in human subjects and animal models of obesity. Also, some studies have shown potential interactions between obesity and aging such as shortening of telomeres, mitochondrial dysfunction, and cellular senescence, and both obesity and aging share some of the common pathways including IGF-1. Most studies have shown that the consumption of anthocyanin-rich food reduces the development of chronic diseases including metabolic diseases and delay the aging process. Tart cherry (TC) (Prunus cerasus) contains the greatest concentrations of anthocyanin flavonoids among vegetables and fruits, including other cherries. However, many studies have used strawberry, blueberry and sweet cherry anthocyanin or extracts to reduce such disease conditions; and fewer studies have addressed the effects of tart cherry on adipose tissue inflammation, obesity and aging. Hence, our objectives were: 1. Identify the potential anti-inflammatory activities of TC extract rich in anthocyanins in adipose tissue inflammation in obese Zucker fatty rats (OZFR) and cultured 3T3-L1 adipocytes; and 2. Identify lifespan regulating effects of TC in C. elegans. Our results of aim 1 showed TC supplementation reduced adipose tissue inflammation both in OZFR and 3T3-L1 adipocytes part via NFkB pathway which is one of the major inflammatory pathways. Moreover, TC increased antioxidant mechanisms in both animal and cell models. In our C. elegans study in aim 2, we found that TC supplementation increased the mean lifespan of wild type C. elegans mainly via IIS pathway. Furthermore, TC increased the expression of antioxidant genes and spare respiration rate in C. elegans. In conclusion, our findings herein demonstrated that dietary TC supplementation may support metabolically healthy obesity through significantly reduced inflammation in adipose tissue. In addition, our work highlighted the importance of TC in delaying aging and improving mechanism involved in healthy aging.Item Role of adipose renin-angiotensin system inhibition and omega-3-fatty acids in obesity and breast cancer crosstalk(2019-12) Rasha, Fahmida Akter; Moustaid-Moussa, Naima; Ramalingam, Latha; Gollahon, Lauren; Rahman, Rakhshanda Layeequr; Kahathuduwa, Chanaka N.Both obesity and breast cancer (BC) are complex diseases of diverse etiologies. Obesity significantly increases the risk for BC; especially in postmenopausal women. Moreover, inflammation is an important underlying basis for both diseases. Thus, it is critical to understand the mechanisms of obesity-related BC and the role of inflammation in linking these diseases. In obesity, several adipocyte-derived inflammatory substances can potentially drive the effects of adipocytes on BC cells. Among these, angiotensin II (Ang II) produced by adipocytes increases local and systemic inflammation in adipose tissue. Ang II is classically known as a hormone that regulates blood pressure and fluid balance. Its unusual production in adipose tissue suggests that it plays an important role in obesity-related hypertension. Various components of the renin angiotensin system (RAS, the system that generates Ang II) are expressed in adipose tissue as well as in BC cells. Interestingly, in obesity, RAS is overexpressed in adipose tissue, leading to increased secretion of Ang II, which is the primary factor behind most RAS-mediated active inflammation. Similarly, overexpression of RAS components drives BC progression via increased cancer cell proliferation, invasion and favorable tumor microenvironment formation. However, limited work has been done to address mechanistic interactions between adipocyte-derived Ang II and obesity-related BC, especially with a focus on modulation of inflammatory pathways by Ang II. On the other hand, given the importance of nutrition in both obesity and BC, it is equally critical to identify nutrients or bioactive food components that can reduce effects of obesity on BC through their anti-inflammatory properties, including reducing RAS-related inflammation in obesity and BC. Omega-3 polyunsaturated fatty acids are among potent anti-inflammatory bioactive compounds known for their anti-obesity and anti-cancer effects. Previous research from our laboratory has shown that part of the anti-inflammatory effects of these fatty acids include reduced secretion of angiotensinogen (Agt, which is Ang II precursor protein in RAS) and other inflammatory cytokines from adipocytes. Accordingly, this dissertation research focused on dissecting the interactions between adipocytes and BC cells via modulation of inflammatory RAS pathway. More specifically, we proposed to (1) determine the role of the adipocyte RAS in BC cell inflammation, proliferation, and invasiveness; (2) determine the role of omega-3 fatty acids alone or combined with RAS inhibitors in reducing effects of adipocytes on BC cells; and (3) identifying potential associations between obesity, antihypertensive medication use (including RAS inhibitors) and BC aggressiveness in BC patients. To accomplish these objectives, we have designed a series of in vitro experiments involving both cultured human or murine mature adipocytes and two types of human BC cells (receptor positive, MCF7 and receptor triple negative MDA-MB-231) to investigate effects of anti-inflammatory RAS inhibitors and omega-3 fatty acids (specifically eicosapentaenoic acid, EPA) on adipocyte-BC cell interactions. In our studies, we attempted to mimic the breast environment in vitro by testing effects of adipocyte-conditioned medium on BC cells. We performed gene and protein analyses along with cell viability and migration assays to determine effects of Ang II, RAS inhibitors and/or EPA either directly on breast cancer cells or using pretreated adipose-conditioned medium (CM). In addition, we also conducted a retrospective pilot study by collecting clinical, pathological and histological data from a convenient sample of female breast cancer patients in collaboration with the Southwest Cancer Center at the Texas Tech Health Science Center to determine associations between body mass index (BMI), antihypertensive medication use (including RAS inhibitors), cancer aggressiveness, and expression of RAS markers in the BC tissues. We demonstrated that direct treatments of BC cells with Ang II, or RAS inhibitors did not alter inflammatory cytokines in either BC cell line. CM from Ang II-pretreated adipocytes significantly increased secretion of pro-inflammatory markers at protein level while RAS inhibitors reduced their secretion in triple negative BC cell line. Additionally, CM from adipocytes treated with RAS inhibitors significantly reduced mRNA levels for markers of inflammation, fat synthesis, and angiogenesis in both BC cell lines. Furthermore, CM from Ang II inhibitor pretreated adipocytes reduced cell migration in both BC cell lines. Similarly, we found CM from human adipocytes pretreated with Ang II inhibitor ± EPA significantly reduced pro-inflammatory cytokines expression and secretion in both BC cell lines, while direct treatments did not alter pro-inflammatory cytokine profile in either BC cells. Moreover, we observed reduced cell migration by triple negative BC cells in response to both direct and CM mediated Ang II inhibitor ± EPA treatments. However, these changes were not significant between individual and combined treatments of Ang II inhibitor and EPA. In the human retrospective pilot study, we demonstrated a positive association between higher BMI with higher cancer aggressiveness, as measured by percent Ki-67 index (marker of cancer cell proliferation). In addition, RAS associated markers including Ang II and its type 1 receptor were expressed in histologic sections of breast cancer tissues. Surprisingly, antihypertensive RAS inhibitor use was positively associated with higher histological grade of BC, which merits further confirmation in a larger cohort of BC patients. In summary, the proposed research is highly significant given the high prevalence of both obesity and BC in women, especially after menopause. One acknowledged limitation is that we did not use adipocytes from healthy control versus breast cancer patients affected with obesity; rather we used mesenchymal stem cells differentiated into adipocytes. This needs to be kept in consideration while interpreting findings from our in vitro studies. We also need to test the mechanistic effect of adipose-Ang II inhibition in multiple breast cancer and control epithelium cell lines for future translational studies. However, our research is novel since it fills an important gap in both pharmacological and nutrition knowledge by repurposing currently used antihypertensive therapies (RAS inhibitors) and anti-inflammatory dietary components (EPA) to attenuate obesity-associated BC. Overall, our findings will aid in identifying new mechanisms to help understand the link between obesity and BC as well as will assist in designing future powered clinical studies with larger patient cohorts.Item Studying the anticancer properties of Parthenolide (PTL) in MCF-7 breast cancer cells(2018-08) Sufian, Hazera Binte; Rahman, Shaikh Mizanoor; Ramalingam, Latha; Khan, ZeinaBackground: Breast cancer is considered as the most prevalent cancer in women world-wide. The average risk for a woman developing breast cancer in her life is about 12% in United States. Cancer cells including breast cancer are resistant to apoptosis and undergo epithelial to mesenchymal transition (EMT) for growth and proliferation. Thus, induction of apoptosis and reversal of EMT process might be effective to prevent cancer cell proliferation and tumor growth. Parthenolide (PTL) is a multifunctional naturally occurring compound. It is considered as a novel anti-tumor agent. Parthenolide induces cytotoxicity in several cancers. However, its anticancer properties in breast cancer cells are not completely known. In this study, low doses of PTL were used to determine its effects on cellular properties, EMT, and angiogenesis in MCF-7 cells and also exploredits underlying molecular mechanism. Methods and materials: MCF-7 breast cancer cells were obtained from American Type Culture Collection (ATCC). Cells were grown in DMEM containing 10% fetal bovine serum and 1% penicillin streptomycin, at 37°C in 5% CO2 incubator. For treating the cells, Parthenolide (PTL) 2 and 5 μM was used for 24 hours using DMEM media with 0.5% FBS. Cell viability was measured using the MTT (3-(4,5-dimethy lthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay kit (BioVision k299-1000). Monolayer wound scratch assay was done for cell migration analysis by using a Nikon Eclipse motorized microscope with incubator (Nikon Instruments Inc.) at 5% CO2 and 37ºC temperature. To detect apoptosis, Apoptosis Detection Kit I (BD Biosciences) was used and the results were analyzed by using a Nikon Eclipse motorized microscope with incubator (Nikon Instruments Inc.). Cytosolic and nuclear extracts were prepared and subjected to Western blot analysis. Proteins and genes implicated in autophagy, EMT and angiogenesis are analyzed by western blot and quantitative polymerase chain reaction (qPCR). For statistical analysis, one-way ANOVA was used. All values are reported as mean ± standard error. Differences were considered to be statistically significant at P values 0.05 or less. Results and Conclusion: Low doses of (2 & 5 µM) of PTL decreases viability, inhibits cell migration, induces apoptosis, and increases autophagy in MCF-7 cells. Interestingly, PTL treatment reverses EMT in MCF7 cells. In addition, PTL activates AMPK as evaluated by identifying phosphorylated form of AMPK. Though we conducted our study in only MCF-7 cells, our results suggest that PTL might be an effective therapeutic option against breast cancer.Item The cellular growth analyzer: a simpler and more comprehensive scratch assay analyzing program(2018-05) Lovelace, Alan; Moussa, Hanna; Kumar, Golden; Ramalingam, LathaA scratch, or wound, assay is a low-cost and simple method to measure cells migration; it is also an easy way to measure the growth rate of cancer cells in vitro. It does so by removing a strip of cells from a cell culture dish and then measuring the cell migration and cellular growth back into the “scratch” area. The measurement of this migration and cellular growth has traditionally been done using the program, ImageJ. Though this method can work, it is labor intensive and time consuming. We therefore developed a more automated and easier-to-use Java program designed specifically to analyze scratch assays. By contrasting and separating the pixels that formed the scratch and then counting them to provide a numerical result the migration and growth we significantly improved the speed and ease of the analysis while allowing the results to be easily repeatable when compared to ImageJ. Further development, refining and addition of new features to this software will significantly aid researchers, especially in the area of cancer research and in assessing the effectiveness of various treatments on cell migration.Item The circulatory levels of irisin in response to acute and chronic resistance training in healthy humans(2015-08) Short, Matthew James; Fernandez del Valle, Maria; McComb, Jacalyn; Ramalingam, LathaObesity is a growing problem in the U.S. and worldwide. Research for potential treatments is abounding. One potential cytokine of interest for the treatment of obesity is irisin. Irisin is released by muscle during physical activity and increases the browning of beige and brown adipocytes. We recruited 31 young and healthy adult males and females (20 female; 11 male) subjects. After blocking and randomly assigning the subjects to either a non-exercise control or exercise intervention group, we analyzed 8 control subjects and 10 exercise intervention subjects after a single bout of high-intensity exercise training. After a 3-week intervention of high intensity resistance training, we analyzed 7 control and 7 exercise intervention subjects. The acute intervention resulted in a significant decreased irisin level during, but not after, an exercise session for the exercise group. The chronic intervention resulted in significant changes in body composition and in an increase in upper and lower body strength in the exercise group. No increase of circulatory irisin was found after 3 weeks of high-intensity exercise training.