bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential Expression of Epidermal Growth Factor Receptor, Hepatocyte Growth Factor Receptor and Hypoxia‐Inducible Factor‐1‐Alpha in Aged Rat Livers
Authors: Etteh Chinedu Casmir , Mee Christopher J
Published: June 2021

Epidermal growth factor receptor (EGFR), hepatocyte growth factor receptor (cMET) and the transcription factor, hypoxia‐inducible factor 1‐ alpha (HIF1‐α) are highly important in angiogenesis, mitogenesis and cell proliferation which are all vital processes required for mammalian liver regeneration. In as much as aging is characterised with general decline in maintenance and function of certain body organs, this is not exactly the same with the liver in regeneration. Aging could result to reduction in the response of the liver to certain extrinsic stimuli such as initiation of stress proteins as a result of injury. To study the effects of age on the expression of these molecules, EGFR, cMET and HIF1‐α gene expression in aged rat liver samples (18months old) vs. young animals (3 months old) was studied through semi and relative quantitative quantification of gene specific mRNA coding for these genes. β‐Actin was used as a reference/housekeeping gene for normalisation of the EGFR, cMET and HIF1‐α genes, the Livak method was used to quantify the relative expression of these genes in relation to the ages of the animals. The results show high mRNA expression of EGFR and cMET in 3 months old samples (young). Analysis of EGFR showed an mRNA fold decrease of 0.56 gene expression and 2.3 for cMET gene express in the 18 months liver. HIF1‐α analysis showed a 5.16 increase in the mRNA gene expression in the 18 month old when compared with the 3 months old. Data from this study show that decrease in expression of epidermal growth factor receptor in the liver with age accelerates decrease in regeneration rate. The fact that EGFR and cMET were expressed at significantly lower levels in older samples reflects the differential decrease in regeneration ability and physiologic activities such proliferation, mitogenesis and angiogenesis in the liver with age. This result in reduction in the ability of the liver to regenerate, thus making the aged liver more susceptible to infections.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigation of Biochemical Effects of Nano‐Silicon Dioxide (SiO2) on Sunflower (Helianthus annuus L.) Plants
Authors: Yurekli Fusun , Kilicaslan Sibel
Published: June 2021

Agricultural biotechnology is very closely related to the properties and potential uses of nanomaterials. Although silicon is not recognized as an important nutrient for plant growth and development, it has beneficial effects on plant growth and yield and on resistance to biotic and abiotic stress. Recent advances in nanotechnology surprisingly increase the use of nanoparticles (NPs) in agriculture; it is therefore interesting to understand the role of SiO2NP. Silicium plays an important role as a physico‐mechanical barrier and is deposited in many plants in the walls of the epidermis, in stem, leaf and vascular tissues, especially in monocots, and also regulates physiological activities in plants. There are also many studies on the regulatory effect of the silicon element on plant growth and development under stress conditions. However, the mechanisms of interaction at the molecular level between the nanoparticles and biological systems are largely unknown. Sunflower seeds were sterilized by washing in 0.5% sodium hypochlorite solution for 5 minutes and then in sterile di‐H2O for 4–5 minutes. Then, on the 1st, 4th and 8th days, samples were taken from root and leaf tissues and frozen in liquid nitrogen after freezing in freezer (−40 ° C). enzyme activity, malondialdehyde (MDA) level, pigment analysis, percentage of germination, relative water content were investigated. In our study, it was determined that 50 nm particle size nanosilicon were more effective at 50 mg / mL concentration. In particular, SOD and APX enzyme activities were found to be highest on the 8th day of nanosilica administration compared to the control group, while CAT enzyme activity was determined in the administration of 100 mg particle size nanosilica 20 mg / mL. MDA levels were decreased on the 8th day compared to the control group. Nanosilica particles help to increase the activity of enzymes that have important roles in combating biotic and abiotic stress conditions and a decrease in MDA levels indicates that nanosilica particles can be taken into account in the cultivation of agriculturally important plants such as sunflower.

Support or Funding Information

This research was funded by Inonu University, Department of Scientific Research Projects Project No: FYL‐2017‐651

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Assessment of bio‐protective potentials of mistletoe leaves extract against biochemical imbalances induced by Combined Oral Contraceptives in rats
Authors: Adebisi Adedayo Adebisi , Sowunmi Tolulope , Ogunlabi Olugbenga Owolabi , Adegbesan Bukunola Oluyemisi
Published: June 2021

Combination birth control pills (Combined Oral Contraceptives ‐ COC) containing female hormones are widely used prescription medications amongst women with up to 70 million users worldwide. This study investigates the effect of prolonged administration of Combined Oral Contraceptives on some biochemical parameters in female wistar rats and the possible bio‐protective potentials of mistletoe leave extract. 30 female wistar rats were divided into three groups of 10 animals each. The groups ‐ control, group E1 and group E2, received daily oral doses of placebo, Nordette® (a COC containing Ethinyl estradiol and levonorgestrel) at 0.03mg/kg body weight and Nordette® + mistletoe leave extract at 600mg/kg body weight respectively for 21 days. Assessment of serum lipid profile, glucose level, ion element homeostasis and total antioxidant status (TAS) of animals were conducted. Our results show that the COC significantly impaired serum lipid profile (p<0.05) (increased total cholesterol, LDL cholesterol, VLDL cholesterol and triacylglycerols, and decreased HDL cholesterol), it increased serum glucose level, it increased TAS and it altered serum ion (Na+, Cl−, K+,Ca2+) levels compared to control animals. There was also a significant (P<0.05) increase in the body and uterus weight of the COC treated rats compared to the control. Co‐treatment of COC with mistletoe leaf extract however protected against these changes. The observed biochemical imbalances in the COC treated rats are common indicators of many health disorders; therefore our result suggests the bio‐protective potentials of mistletoe leaf extract against these physiological insults.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The influence of neonatal intake of curcumin on expression of genes associated with lipid metabolism and inflammatory cytokines: Implication on obesity
Authors: Mukwevho Emmanuel , Matumba Mashudu Given
Published: June 2021

The high prevalence of obesity is a serious global threat to the public's health and at least 41 million children younger than five years are overweight or obese, most of whom live in low‐income and middle‐income countries. Furthermore, obesity is a major risk factor for the development of type‐2 diabetes, cardiovascular disease, cancer, disability, hypertension and stroke. This current study investigated the effect of curcumin on expression of genes associated with lipid metabolism (AMPK, ACC‐1, CPT‐1, SREBP‐1, and PGC‐1) since they are known to be impaired in obesity situations. The study also investigated the effect of curcumin on inflammatory cytokines (TNF‐α, and IL‐6). This was motivated by the report indicating that obesity is associated with chronic inflammation, given evidence by the increased levels of cytokines in adipose and liver tissues. Curcumin is one the hundreds of components isolated from ancient spice turmeric and it is being studied intensively today for its potential therapeutic activity in the treatment of inflammatory diseases such as obesity, diabetes and cardiovascular diseases Therefore to achieve our aim, Sprague Dawley rats were fed with high fructose diet to induce obesity model, and the rats that developed metabolic syndrome were considered as diseased, they were then treated curcumin. Analysis of gene expression was done in liver tissues using Real‐Time PCR technique. Results clearly indicated that curcumin significantly improved all the genes associated with lipid metabolism in rats that were treated with curcumin. Furthermore, results showed significant increase on all inflammatory cytokine gene expression of rats that were induced with metabolic syndrome. These results demonstrate that curcumin produce good effects in ameliorating neonatal metabolic syndrome since it was able to downregulate all the inflammatory gene expression and improved genes associated with lipid metabolism. In conclusion, these study confirms that curcumin plays a key role on lipid metabolism and inflammatory cytokine regulation and it can be used as an effective therapeutic agent to ameliorate neonatal metabolic syndrome.

Support or Funding Information

Funding is from National Research Foundation (NRF) and North‐West University.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hexokinase activity were decreased in erythrocytes and increased in blood serum at the DM
Authors: Shamansurova Zulaykho , Saatov Talat , Akhmedova Mukhabbat
Published: June 2021

Background

Diabetes Mellitus (DM) are combination of metabolic abnormalities due to chronic hyperglycemia state, which is reflects from cells to whole body. Hexokinase as a ubiquitous enzyme important in catch and clamp glucose from outer space inside of cells. Regulation of hexokinase activity would help to restore the glycemic control at the DM.

With aim of determining hexokinase activity in erythrocytes and in blood serum 42 patients with type 2 DM were observed. Hexokinase activity calculated by colorimetric enzyme‐substrate reaction. In addition the fasting blood glucose, HbA1c, plasma sialic acids level and Erythrocytes sialidase activity were measured in this patients.

Results shown that hexokinase activity were decreased in erythrocytes (78%, P<0.05) and increased in blood serum (115%, P<0.01), where the strong linkage between decreasing of erythrocytes hexokinase activity and increasing in the blood serum with fasting plasma glucose and HbA1c level were detected. Erythrocytes sialidase activity were increased (134%, P<0.01) and blood plasma sialic acid level were increased (154%, P<0.01) in patients with DM and indicated the degree of the structural and functional abnormalities on the cell surface. The strong linkage between erythrocytes sialidase activity and blood plasma sialic acids level were detected with blood plasma glucose and HbA1c level.

Conclusion

The hexokinase activity were decreased in erythrocytes and increased in blood serum in patients with DM in depended with blood glucose and HbA1c and can be used as a marker of the cell membranes structural and functional abnormalities.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pre‐Transplant Medical Arterial Calcification and Post‐Transplant Hypertension
Authors: Tantisattamo Ekamol , Vutthikraivit Possawat , Dejhansathit Siroj
Published: June 2021

Background

Medial arterial calcification (MAC) is common in chronic kidney disease (CKD) and end‐stage renal disease (ESRD). Breast arterial calcification detected by mammogram (MMG) is exclusively medial and can lead to arterial stiffness. The association between pre‐transplant MAC and hypertension (HTN) after kidney transplantation (KTx) is unknown.

Method

MMG performed closest to the date of KTx were examined for presence or absence of MAC. The association between MAC and systolic hypertension (SHTN) defined by systolic blood pressure (SBP) ≥130 mmHg within the first 48 weeks after KTx is examined as time‐to‐event by Cox proportional hazard analysis.

Results

Of all 51 kidney transplant recipients from a single kidney transplant center followed up 48 weeks after KTx, mean±SD age was 57.08±10.47 years. Ninty‐six percent of patients had pre‐transplant HTN and 41% had diabetes. Mean BP was 143.57±25.77 / 76.61±13.75 mmHg. Mean±SEM pre‐transplant SBP in patient without pre‐KTx MAC was lower than those with pre‐transplant MAC (140.74±4.65 VS. 147.95±5.73; 95% confidence interval (CI) −22.07, 7.65; p 0.3344). Compared to the patients without MAC, MAC patients had 31% greater risk of having SHTN within 48 weeks post‐KTx (HR 1.31, 95%CI 0.73, 2.37, p 0.367; Figure 1). After adjusted for age, cause of ESRD, type of KTx, type of induction immunosuppressive medications, presence or absence of pre‐KTx HTN, diabetes, BMI, DBP, and corrected calcium, the association was reversed but still not statistically significance (HR 0.72, 95%CI 0.34, 1.50, p 0.374).

Conclusion

Although MAC is associated with arterial stiffness in non‐transplant patients, no association between presence of pre‐transplant MAC and SHTN may indicate change in vascular stiffness. Successful KTx may modify factors effecting vascular milieu associated with MAC.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigation of Some Angiogenic Factors Related to Aging in Brown and White Adipose Tissues of Rats
Authors: Yurekli Muhittin
Published: June 2021

Aging is a biological phenomenon in which life force decreases, there is an increase in dysfunction, and activities at the level of tissue and cell with insufficient response to changes in internal and external environments decrease. In biological aging, there are deficiencies in the cell level, the cells and tissues are prevented from feeding with enough oxygen and hypoxic conditions can occur. Due to the loss of function in living organisms as a result of aging, the failure of the organelles to perform their functions may lead to the emergence of oxygen deficiency conditions in the tissues. The most common feature of aging is the change of physiological functions over time. Especially when oxygen deficiency appears in the tissues, the number of red blood cells increases, hormones and angiogenic factors releasing increase. Hypoxia may also occur in tissues due to environmental factors, obesity and various circulatory insufficiency, and the organism may activate angiogenic factors in response to this. Adequate oxygen must be provided for the normal functioning of the tissues. In the case of oxygen deficiency (hypoxia), matrix metallo proteinase, vascular endothelial growth factor, tumor necrosis factor, interleukin may play role such as angiogenic factors. In this study, the levels of vascular endothelial growth factor (VEGF), matrix metalloproteinase‐II, hypoxia‐inducible factor‐1α, adrenomedullin were investigated in 6 months and 24 months old rats. The vascularity function of the tissues is very important. Vein formation is required in both normal functional tissues and in tissues that are in neoplastic formation. Therefore, in this study, we investigated the angiogenic factors in rats' brown and white adipose tissue in cases by aging. In the study, 24 rats were divided into two groups; 12, 6‐month old young and 12, 24‐month‐old rats were evaluated as elderly group. In the study, angiogenic factors increased in the white adipose tissues of the elderly animals, whereas it was found that there was a greater increase in the brown adipose tissue that plays a role in the thermogenesis event in young rats. Decreasing physical activity in elderly animals and increased activity in white adipose tissue resulted in a significant increase in the weight of aged rats. Angiogenic factors play an important role in maintaining the oxygen demand of the organism in elderly animals, while it may cause an increase in the expanding of the white adipose tissue, which is effective on weight gain, which may lead to obesity.

Support or Funding Information

This research was funded by Inonu University, Department of Scientific Research Projects Project No: FYL‐2017‐748).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Breast cancer cell invasiveness is stimulated by loss of membrane interaction of actinbinding protein profilin1 via altered phosphoinositide metabolism
Authors: Roy Partha , Gau David , Bae Yongho , Ohayon Lee
Published: June 2021

Profilin1 (Pfn1) belongs to a class of actin‐binding protein cytoskeletal function of which is negatively regulated by its interaction with membrane phosphoinositides (PPI). In stark contrast to most physiological scenarios where loss of Pfn1 significantly impairs normal cell migration and invasion through inhibition of actin polymerization, breast cancer cells exhibit hyper‐migratory phenotype in vitro and enhanced hematogenous dissemination from primary tumor in vivo upon depletion of Pfn1, the underlying mechanism of which is not easily explained by Pfn1's cytoskeletal function. Through knockdown and overexpression studies in multiple cell lines, we show that Pfn1 plays a key role in determining the cellular profile of various PPIs including PI(4,5)P2, PIP3 and PI(3,4)P2 Specifically, Pfn1 promotes PI(4,5)P2 and inhibits membrane accumulation of PI3K‐generated PPIs including PIP3 and PI(3,4)P2, a feature related to Pfn1's PPI interaction. Kinetics of PI3K signaling output suggest that loss of Pfn1 enhances sustenance of PI3K signaling by inhibiting an antagonistic pathway, a finding that correlates to Pfn1‐dependent regulation of PTEN. We establish Pfn1‐dependent alteration of multiple pro‐metastatic cytokines and membrane recruitment of selective PI(3,4)P2‐dependent pro‐migratory protein complexes, the latter in particular linking to hyper‐migratory and hyper‐invasive phenotypes of Pfn1‐deficient breast cancer cells. In summary, these findings establish novel regulation of cell migration by lipid signaling via membrane interaction of actin‐binding protein.

Support or Funding Information

NIH 2R01CA 108607

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Fatty Acids Bind to Them1, a Negative Regulator of Thermogenesis in Brown Adipose Tissue
Authors: Tillman Matthew C , Khadka Manoj , Okafor Chiamaka Denise , Juneja Puneet C , Ortlund Eric A
Published: June 2021

Obesity and other metabolic diseases have become a global epidemic, greatly increasing in prevalence over the past 30 years. In mammals, brown adipose tissue counteracts obesity through thermogenesis: the burning of fat to produce heat. Thioesterase superfamily member 1 (Them1) is a multi‐domain enzyme in brown adipose tissue that slows down thermogenesis by converting the substrate acyl‐CoA into fatty acids. Additionally, Them1 contains a lipid‐binding StAR‐related lipid transfer (START) domain with an unknown function. We hypothesized that the lipid‐binding START domain controls thermogenesis by functioning as a lipid sensor for the enzymatic domains of Them1. We used mass spectrometry, X‐ray crystallography, and microscale thermophoresis to demonstrate that long‐chained fatty acids (14–20 carbon tail length), products of Them1's enzymatic reaction, bind to the START domain. Additionally, we determined a low‐resolution structure of full‐length Them1 using negative stain electron microscopy and found that Them1 forms a trimer, making contacts between the enzymatic domains. Using this structure, we ran molecular dynamic simulations to compare protein dynamics in the presence and absence of fatty acid. We determined that the largest differences in protein motions were localized to the START domain. In conclusion, we have used a robust, multidisciplinary approach to demonstrate that fatty acids, the product of Them1 enzymatic activity, bind the START domain, making Them1 the first protein of the START domain family identified to bind fatty acids. Additionally, we provide the first preliminary structure of full‐length Them1, which enhances our limited knowledge of multi‐domain thioesterases.

Support or Funding Information

This study was supported in part by the Emory Integrated Lipidomics Core (EILC), which is subsidized by the Emory University School of Medicine and is one of the Emory Integrated Core Facilities. Additional support was provided by the Georgia Clinical & Translational Science Alliance of the National Institutes of Health under Award Number UL1TR002378. The content is solely the responsibility of the authors and does not necessarily reflect the official views of the National Institutes of Health.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MEK1/2 inhibitor reduces vascular calcification by regulating both canonical and non‐canonical Wnt signaling pathways
Authors: Zeng Peng , Yang Jie , Liu Lipei , Chen Yuanli , Han Jihong
Published: June 2021

We previously reported that MEK1/2 inhibitors, such as PD98059 or U0126, synergized liver X receptor (LXR) ligand, such as T0901317, activated macrophage ATP‐binding cassette transporter A1 (ABCA1) expression and free cholesterol efflux. Therefore, the combined U0126 and T0901317 can regress the established lesions. Additionally, U0126 increased lesion stability with unknown mechanisms. Vascular calcification is a major risk factor for plaque rupture. In this study, we determined if MEK1/2 inhibitors can affect atherosclerotic calcification and the underlying mechanisms. In vivo, we determined that treatment of apoE−/− mice with U0126 reduced HFD‐induced atherosclerotic calcification. It also reduced calcification medium‐induced calcium deposition in aortic rings ex vivo or in human aortic smooth muscle cells (HASMCs). Associated with reduction of vascular calcification, expression of osteogenic genes, such as bone morphogenetic protein 2 (BMP2), runt‐related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), β‐catenin and osteopontin (OPN), were reduced by U0126, either in vivo or in vitro. Mechanistically, MEK1/2 inhibitors increased dickkopf 1 (DKK1) secretion and LDL receptor‐related protein 6 (LRP6) expression, thereby inhibiting canonical Wnt/β‐catenin signaling pathway while activating LRP6‐induced non‐canonical Wnt signaling pathway. In contrast, inactivation of either DKK1 or LRP6 attenuated the anti‐vascular calcification effects of MEK1/2 inhibitors. Based on the findings above, we conclude that MEK1/2 inhibitor can rescue vascular calcification through regulation of both canonical and non‐canonical Wnt signaling pathways. Our study also suggests inactivation of MER1/2 by chemicals can be a potential therapeutic strategy for treatment of atherosclerosis and atherosclerotic calcification.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Targeting Molecular Chaperone HSP90 To Treat Niemann‐Pick Type C1 Disease
Authors: Pipalia Nina H , Subramanian Kangaraj , Cross Abigail , Garg Kunal , Al‐Motawa Aisha , Maxfield Frederick R
Published: June 2021

NPC1 is a fatal, neurodegenerative lysosomal storage disease caused by mutations in the NPC1 gene. More than 300 different disease‐associated mutations are reported in patients, resulting in abnormal accumulation of unesterified cholesterol, glycosphingolipids and other lipids in late endosomes/lysosomes (LE/Ly) of many cell types. There is no FDA‐approved therapy to treat NPC1 disease. A limited initial human trial of hydroxylpropyl beta cyclodextrin (HPBCD) is currently in progress. Our previous studies suggested that treatment with histone deacetylase inhibitors (HDACis) on skin fibroblasts results in increased expression of the NPC1 protein in mutant cells. Using an engineered human cell line, we have observed that HDACis can correct the NPC1 cholesterol storage defect in 60 of the 81 NPC1 mutants tested. However, when HDACis', were tested on non‐dividing cells like bone marrow derived macrophages, we did not observe any correction.

Although, significant progress has been made in understanding the pathology of NPC1 disease, the mechanisms governing the proteostatic control of NPC1 protein is not well understood.. Disruption of molecular chaperone HSP90 (via hyperacetylation) is one of the many consequences of HDACi treatment. Thus, we evaluated the effect of direct inhibition of HSP90 family of proteins on NPC1 skin fibroblasts in terms of lowering lysosomal cholesterol accumulation.

The data on the effect of several different HSP90 inhibitors on human patient‐derived skin fibroblast in lowering lysosomal cholesterol burden using microscopy based filipin assay will be presented. Our results suggests that cytosolic HSP90 and associated chaperones may be a target and that protein folding quality control plays a critical role in NPC1 diseases. We observed that out of the three HSP90 family proteins, inhibiting HSP90A and not HSP90B or TRAP resulted in clearance of cholesterol from lysosomes as well as aided in mobilizing unfolded mutant NPC1 protein from ER to lysosomes. We will also present the results elucidating the role of cytosolic HSP90 and its associated co‐chaperones in the regulation of NPC1 mutant protein stability.

Support or Funding Information

1. MDBR‐Orphan Disease UPenn Grant to Dr. Pipalia 2. APMRF Grant to Dr. Maxfield3. DART Summer Fellowship to Abigail Cross

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Vitamin E Reduces Hepatic VLDL Secretion by Disrupting its ER‐to‐Golgi Transport
Authors: Siddiqi Shadab A , Clay Ryan , Siddiqi Shaila
Published: June 2021

Increased secretion of triglycerides rich‐very low‐density lipoproteins (VLDL) by the liver causes hyperlipidemia, which is associated with a variety of metabolic disorders. The rate‐limiting step in the secretion of VLDL particles from the liver is their transport from the endoplasmic reticulum (ER) to the Golgi and represents a potential therapeutic target in controlling VLDL secretion. Our laboratory has demonstrated that the targeted delivery of VLDL from the ER to the Golgi is facilitated by a distinct ER‐derived vesicle, VLDL transport vesicle (VTV). In this study, we investigated the role of vitamin E (alpha‐tocopherol) in the intracellular VLDL trafficking and secretion. To determine the role of vitamin E, we treated primary hepatocytes with different concentrations of vitamin E complexed with bovine serum albumin (BSA), carried out VTV‐biogenesis assays to examine ER‐to‐Golgi transport of VLDL and determined VLDL secretion. Our data demonstrate that vitamin E‐treatment of hepatocytes significantly reduces VTV‐biogenesis from the ER membranes and VLDL secretion. Interestingly, we found that vitamin E‐treatment significantly reduces the expression of proteins, Sar1b and SVIP, that control ER‐to‐Golgi transport of VLDL. We conclude that vitamin E controls VTV biogenesis by regulating the expression of Sar1b and SVIP proteins and thus plays an important role in VLDL secretion by the liver. This study was supported by NIDDK, NIH R01 DK081413 to SAS.

Support or Funding Information

This study was supported by NIDDK, NIH R01 DK081413 to SAS.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Lysosome‐Associated Membrane Protein 1 Rescues Cholesterol Accumulation in NPC1 Mutant Cell by Enhancing Lysosome‐ER Contacts
Authors: Singhal Ashutosh , Song Byeongwoon
Published: June 2021

Niemann‐Pick type C (NPC) disease is a neurodegenerative disorder caused by mutations in NPC1 or NPC2 gene, encoding a lysosomal membrane and luminal protein, respectively. NPC disease is linked to the disruption of intracellular cholesterol trafficking and is characterized by excessive accumulation of free cholesterol in lysosomes. The functions of NPC1 and NPC2 proteins are involved in the exit of lysosomal cholesterol by poorly understood mechanisms. Understanding the molecular mechanisms involved in lysosomal cholesterol export is critical for NPC therapy.

We hypothesized that the lysosome‐associated membrane protein 1 (LAMP‐1) can mediate lysosomal cholesterol export in the absence of NPC1 function. By filipin staining, we demonstrated that 2‐hydroxypropyl‐β‐cyclodextrin (HPβCD) or HPγCD treatment rescues lysosomal cholesterol accumulation in NPC1 mutant cells derived from NPC patient. By proteomic analysis, immunoblotting, and confocal microscopy, we showed that HPβCD or HPγCD treatment in NPC1 mutant cells increases the expression of the LAMP‐1 protein and redistributes lysosomes toward the cell periphery. Overexpression and knockdown of LAMP‐1 in HeLa cells suggested that LAMP‐1 plays an important role in lysosomal cholesterol export. By confocal microscopy, we further demonstrated that LAMP‐1 is involved in establishing membrane contacts between lysosomes and the endoplasmic reticulum (ER). By analyzing the cholesterol content and proteome profile of isolated lysosomes, we provide new insights on the effect of HPβCD and HPγCD in modulating lysosomal functions and in reducing lysosomal cholesterol accumulation in NPC1 mutant cells.

In conclusion, our studies suggest that LAMP‐1 can restore cellular cholesterol homeostasis by modulating lysosome positioning, potentially enhancing lysosome‐to‐ER cholesterol transport for cholesterol esterification and metabolism. Further studies on LAMP‐1 functions and lysosome dynamics are warranted to better understand NPC disease mechanisms and to design effective therapies for NPC.

Support or Funding Information

This study was supported by the Startup Fund, Pilot Grant, and Seed Grant to B.S. from Meharry Medical College.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evaluation of Dietary Protein Isolated from Different Sources on Hepatic and Adipose Lipid Metabolism
Authors: Warren Derek Cole , Skinner Roy Chris , Laub David , Barnes Kimbery , Matak Kristen , Tou Janet C
Published: June 2021

Dietary protein is a significant contributor to the process of lipotoxicity through the modulation of insulin secretion and the regulation of adipocyte metabolic function. However, the contribution of different types of dietary protein is less understood. Previous studies have reported the effects of proteins isolated from multiple animal sources on liver lipid metabolism did not differ from casein; therefore, the objective of this study was to evaluate lipid metabolism in the liver and adipose in rats fed protein isolated from aquatic sources. Young (age 28 days) female Sprague‐Dawley rats were randomly assigned (n=8 rats/group) to standard purified diet with 10% w/w protein as casein, whey, Silver carp isolate or a protein‐free diet. There were no significant differences in liver weight or lipid content among rats fed these different protein sources. However, the protein‐free group had the highest hepatic lipid content (19.96% ± 4.13) among the diet groups confirmed by histological evaluation liver showing greater fat infiltration. Liver fatty analysis showed that rats on a protein‐free diet had the lowest (p<0.05) stearic acid content (6.77% ± 0.43), and the highest (p<0.05) oleic acid, linoleic acid, and α‐linolenic acid content at 20.81% ± 1.29, 23.21% ± 1.67, 1.56% ± 0.09, respectively. There were no differences in fatty acid composition in the liver of rats fed casein, whey, or carp isolate. However, rats fed whey isolate had greater (p<0.05) body weight and gonadal fat pad mass but no significant differences gonadal adipose fatty acid composition or serum LDL, HDL, and cholesterol compared to rats fed casein or carp. Collectively, these results indicated the amount rather than dietary protein source influences adipose tissue and hepatic lipid metabolism.

Support or Funding Information

This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2015‐67017‐23084 from the USDA National Institute of Food and Agriculture

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Targeted Interleukin‐27 Gene Therapy Development for Disrupting Key Malignant Crosstalk between Metastatic Prostate Cancer and Bone
Authors: Salameh Janelle W. , Figueiredo Marxa L.
Published: June 2021

Prostate cancer (PCa) bone metastases have been reported in up to 90% of patients with advanced disease. Prostate cancer cells ‘hijack’ bone homeostasis by disrupting the crosstalk between critical cells within the tumor/bone microenvironment (osteoblasts, osteoclasts, and immune cells), and utilizing this effector‐rich environment for cancer cell survival and growth. Crosstalk pathways primarily involve Interleukins (IL‐6 and IL‐11) and transforming growth factor β (TGFβ). Therefore, a key therapeutic objective in malignant skeletal disease management is to eliminate tumors while restoring bone homeostasis. Current treatments are limited to palliative radiotherapy, chemotherapy, or anti‐RANK treatments, all of which have considerable side effects such as osteonecrosis of the jaw or enhanced tumor invasion. An emerging approach to treating skeletal malignancies, osteoimmunology, investigates new multifunctional immune‐stimulatory agents that can simultaneously combat tumor growth, and promote bone regeneration. IL‐27 is an immunomodulatory cytokine with a great potential as a multifunctional cancer therapeutic. Its regulatory roles include antitumor immune responses (promoting Th1 differentiation), inhibition of tumor growth, and direct modification of tumor/bone crosstalk to support bone regeneration. Thus, we hypothesize that a tumor‐targeted IL‐27 gene therapy approach would facilitate the accumulation of IL‐27 in the bone/tumor microenvironment and enhance its therapeutic efficacy. To test this hypothesis, we created a tumor‐targeted IL‐27 gene by incorporating a novel peptide (LSLITRL); an IL‐6 receptor antagonist known to inhibit angiogenesis and tumor growth; or a scrambled control peptide at the C‐terminus. To detect the expression, secretion, and tumor recognition specificity, IL‐27 constructs were fused to a NanoLuciferase expressing vector. We have also assessed the potential for tumor‐targeted IL‐27 expression in disrupting malignant characteristics of PCa in vitro in terms of viability and invasiveness following transfection. Future studies will examine the expression and therapeutic efficacy of tumor‐targeted IL‐27 in vivo in a TRAMP‐C2 tibial implantation model.

Support or Funding Information

NIH‐NCI R01CA196947‐S1

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Powerhouse of Depression: The Impact of MTCH‐1 Deficiency on Reward Signaling in C. Elegans
Authors: Penchina Kaitlyn , Zuklie Angus , Adams Clay , Dittmar Caleb , Park Madeline , Fong Megan , Langner Grace , Cole Austin , Madejski Sophia , Sweeney Robert , De Luke
Published: June 2021

Major Depressive Disorder (MDD) is the leading cause of disability worldwide. One of MDD's most troubling symptoms is the loss of reward firing, or anhedonia. Current publications indicate that MTCH‐2, a gene associated with mitochondrial transport, is related to a decrease in firing rates in the hippocampus of mice. Additionally, a GWAS reported a link between MTCH‐2 and neuroticism in humans. Consequently, we hypothesize that a MTCH‐2 related decrease in firing rates in the nucleus accumbens could potentially play a role in the development of depressive symptoms.

The C. Elegans MTCH‐1 protein shows a 87% homology to the human MTCH‐2 protein. Here, we examine the relationship between the MTCH‐1 gene and C. Elegans reward firing. Because C. Elegans reward firing levels have been associated with changes in speed change fluidity in past studies, we measure C. Elegans motility as a proxy for reward firing. In this study, we compare motility and gene expression in MTCH‐1 deficient worms and wt worms and further examine MTCH‐1 knockdown to control motility. Together, these data points may indicate a role of MTCH‐2 in depression.

Support or Funding Information

The Nueva School

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Morpho‐structural Peculiarities of Blood Formed Elements and Clinic‐pathological Characteristics in Women with Uterine (Benign, Malignant) Tumors in Menopausal Age
Authors: Kotrikadze Nanuli , Tufinashvili Tamar , Nakashidze Irina , Ramishvili Liana , Alibegashvili Manana , Gordeziani Manana , Sepiashvilli Bela , Ahmad Sarfraz
Published: June 2021

Objective

To evaluate ongoing alterations that reflect on the structural characteristics of blood formed elements based on the hormonal imbalance and to investigate the ABO system antigens among menopausal women with uterine (benign & malignant) tumors.

Methods

Blood specimens from women with benign (n=15), malignant (n=15) uterine tumors, & healthy menopausal women (control, n=15) were used. ELISA kits were used for the quantitative determination of hormones. The blood formed elements ultrastructure observations were conducted by electronic microscope.

Results

Compared to control (33.8±0.7 pg/mL), estradiol level was higher in benign (45.7±0.9 pg/mL) & malignant (70.7±3.7 pg/mL) cases (p<0.001). Similar pattern was noted in testosterone levels [control=0.38±0.03 ng/mL, benign=0.55±0.04 ng/mL (p<0.01), malignant=1.56±0.14 ng/mL (p<0.001)] was higher in malignant cases. In contrast, progesterone levels were decreased in the disease cases [control=0.93±0.05 ng/mL, benign=0.44±0.003 ng/mL, 0.31±0.02 ng/ml (p<0.001)]. The percent ABO system antigens distributions were: O(I) (control=60±10.9, benign=40±10.2, malignant=25±10.2), A(II) (control=30±10.2, benign=55±11.1, malignant=70±10.6), B(III) (control=5±4.8, benign=0±0, malignant=0±0), and AB(IV) (control=5±4.8, benign=5±4.8, malignant=5±4.8) percent. Assessments of morphologic structure of erythrocytes revealed numerous pathological form erythrocytes (Poikilocytosis) as benign, also in case of malignant tumors, particularly target cell (codocytes), hamlet cell, Teardrop cells (dacrocytes), Sickle cells (Drepanocytes) erythrocytes.

Conclusions

Using ELISA and transmembrane electron microscopy, our results demonstrate that in the malignant uterine tumor of menopausal women, quantitative/structural changes occur in blood formed elements indicating ongoing alterations in hormonal imbalance. Assessing these changes in structural characteristics would be useful in examining uterine pathologies and subsequent treatment plans.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Combination of Ionizing Radiation with Glutaminase Inhibition Improves Treatment Response in Head and Neck Squamous Cell Carcinoma
Authors: Wicker Christina Ann , Mills Gordon , Wise‐Draper Trisha , Takiar Vinita
Published: June 2021

Head and neck cancer is the 6th most common cancer worldwide. Ionizing radiation (IR) is an important aspect of cancer treatment, however its efficacy is limited by radioresistance, resulting in locoregional failure rates as high as 50%. The pathways used by cells to evade the cellular damage caused by IR are poorly understood. Initial Reverse Phase Protein Array analyses in our laboratory demonstrated that IR induced metabolic pathways, including those involving the TCA cycle. Glutamate is a key substrate of the TCA cycle, and essential for the production of biomolecules necessary for tumor proliferation. Glutaminase is the enzyme that converts glutamine to glutamate. Analysis of The Cancer Genome Atlas's transcriptome database revealed that glutaminase overexpression (upper 25%) in patients with head and neck squamous cell cancer (HNSCC) was associated with significantly reduced patient survival (p<0.03) . In this study, we aimed to identify whether glutaminase inhibition with CB‐839 (Caymen Chemical) could enhance the cellular response to IR. To do this, we used three representative human‐derived HNSCC cell lines: CAL‐27, FaDu, and HN5. We first established that proliferation of all three HNSCC cell lines was glutamine dependent by Dojindo CCK‐8 cell proliferation assay (20–50% increase, n=3 per line, p<0.01). Moreover, a synergistic decrease in cell survival was observed in clonogenic assays with HNSCC cell lines treated with IR and CB‐839 relative to either treatment alone (n=3 per line, p<0.05). In 3D cell culture, which better represents the host microenvironment, combinatorial treatment significantly reduced HNSCC spheroid size (11–26% decrease, n=3 per line, p<0.0001). To further understand the mechanism behind this reduced cell growth and survival, we examined CB‐839's effect on cellular metabolism using Seahorse MitoStress test assays. CB‐839 significantly reduced spare respiratory capacity and OCR/ECAR rate in HNSCC cell lines examined (p<0.001, p< 2.0×10−6,n>3). Mims et al. 2015 previously demonstrated that suppressing spare respiratory capacity enhances radiation sensitivity, which may explain the observed synergistic effects of CB‐839 in combination with IR. In summary, radiation resistance and locoregional failure is a major hurdle in the successful treatment of HNSCC. Combinatorial treatment with IR and the glutaminase inhibitor, CB‐839, led to an enhanced response over IR alone in HNSCC. These results support glutaminase as a possible target to overcome radiation related treatment resistance in patients with HNSCC.

Support or Funding Information

Pathways to Cancer Therapeutics Training Grant (CMP & SW T32CA117846‐11A1)

VA Career Development Award (V.T. IK2 BX004360‐01A1)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cell Strains Derived from Long‐term 3D Tumor Models of MCF‐7 Display Characteristics and Phenotypes Consistent with Stem‐cell Populations
Authors: Topel Samantha , Lyden Timothy
Published: June 2021

In a continuing series of studies, our laboratory has been examining the cell populations associated with long‐term 3‐D artificial tumor models generated using standard MCF‐7 breast ductal adenocarcinoma cells. In these studies we have developed long‐term cultures and grown them out to 1, 2, 3 and 4 years respectively. This report focuses on MCF‐7 tumors developed on de‐cellularized and de‐calcified bovine trabecular bone scaffolds. These long‐term cultures generate populations of cells throughout the entire culture period by shedding both individual cells and cell clusters, often in the form of spheroids. These various shed cell populations then form colonies and eventually monolayers of cells, from which the scaffold must be removed to new wells as a form of “passage”. This entire process of monolayer formation then repeats itself again within a week in the new wells. In addition to shed cells we have also harvested cell strains directly from the scaffolds by standard trypsinization for 15 minutes followed by a second trypsinization of an addition 15 minutes. The paired cultures of rapidly released and slowly released cells display significantly different phenotypes and present with differential marker patterns as well. In addition, we have also established cell strains from both explanting cells directly associated with the artificial tumors and colonies which establish themselves at a distance from the original tumor. These distinct populations also display unique phenotypes and marker patterns by flow cytometry. In these analyses it is clear that at least two major populations occur in both types of monolayer, but that the relative proportion is distinctly different, with a much increased stem‐like cell population being observed in released and floating cells from the colony strain but far less so from the explanting strain. Based on both size and CD44/CD24 immunophenotyping, we have established that this distinct cell population is mainly associated with distant colonies. It is our working hypothesis that these respective populations represent differential metastatic states in artificial tumor‐derived cell strains. Furthermore, we contend that this work characterizing cell populations derived from long‐term tumor models further confirms our overall conclusion that these artificial tumor tissues provide an effective model of tumor development including progression and metastasis in‐vitro.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Choline acetyltransferase: A novel molecular target in lung adenocarcinoma therapy
Authors: Richbart Stephen D , Akers Austin T , Friedman Jamie R , Bow Eric W , Rimoldi John M , Hardman Elaine W. , Dasgupta Piyali
Published: June 2021

The clinicopathological properties of lung adenocarcinoma (LAC) in smokers is divergent from LAC in non‐smokers. One of the distinctive features of LAC in smokers is that the disease is relatively resistant to targeted molecular therapies. For example, targeted therapeutic agents like EGFR‐inhibitors (erlotinib and gefitinib) are highly effective in LACs in non‐smokers. However, these agents display much lower anti‐tumor activity in LACs in patients who are active smokers. Similarly, lung cancer patients (who are active smokers) show a lower response to chemotherapy than those who are non‐smokers. However, the majority of LAC patients are smokers. This underlines the need to identify viable drug targets for LAC therapy in patients who are exposed to cigarette smoke. A survey of literature reveals that nicotine (the addictive component of cigarette smoke) accelerates the growth of lung cancers, as well as confers resistance to chemotherapy. One of the mechanisms underlying the mitogenic activity of nicotine is that it promotes the production of the neurotransmitter acetylcholine (ACh) from LAC cells. ACh is known to be an autocrine growth factor for LAC cells and is synthesized by the enzyme choline acetyltransferase (ChAT). The present study investigates the feasibility of ChAT as a molecular target for LAC in smokers. We find that ChAT levels are upregulated in human LAC cell lines and tissues in analogous to their smoking history. Finally, the ChAT inhibitor BW813U causes robust apoptosis in human LAC cell lined and LAC cell lines isolated from patients. The magnitude of BW813U‐induced apoptosis is similar across LAC cell lines irrespective of smoking history; however, the concentration of BW813U which causes apoptosis is lower in LAC cell lines belonging to heavy smokers. The anti‐tumor activity of BW813U is also observed in H838 cells (belonging to an 80 pack‐year smoker) xenografted on athymic mice. Our studies show that antagonists of ChAT like BW813U may have therapeutic applications in the majority of the population of LAC who are smokers.

Support or Funding Information

Funding for our study was supported by a NIH R15‐AREA Grant (2R15CA161491‐02). Furthermore, this study was supported in part by an Institutional Development Award (IDeA) Grant number P20GM104932 from the National Institute of General Medical Sciences (NIGMS) and the Research Core B of COBRE, a component of the National Institutes of Health (NIH).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.