bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intermittent Hypoxia Upregulates Angiotensin II Generating Pathways in Human Preadipocytes
Authors: Becari Christiane , Singh Prachi , Polonis Katarzyna , Somers Virend K. 
Published: November 2023

BackgroundObstructive sleep apnea (OSA) is characterized by repetitive intermittent hypoxia and reoxygenation during sleep, and is a risk factor for cardiometabolic diseases including hypertension and diabetes. The renin‐angiotensin system (RAS) is an important cardiovascular and metabolic regulator playing key roles via angiotensin peptides. Recent studies demonstrate that all components of the RAS, including enzymes responsible for AngII generation, such as angiotensin converting enzyme (ACE) and chymase, are present in adipose tissue. Furthermore adipose tissue significantly contributes to systemic levels of AngII, thereby potentiating development of hypertension in obesity. Given that OSA patients are mostly obese, the localized effects of RAS in adipose tissue are likely an important mediator of OSA‐related hypertension. This study sought to investigate the effects of intermittent hypoxia (IH), a hallmark of OSA, on Ang II generation pathways in preadipocytes.Methods/ResultsRAS plays an important role in adipose tissue via its action on preadipocytes. Therefore, we examined the effects of IH in primary cultures of human white preadipocytes (HWP). HWP cells were exposed to repeated cycles of 30 min‐21% O2 (normoxic) and 30 min‐1% O2 (hypoxic) at 37°C for 24 hours. Cells grown in continuous normoxic conditions were used as controls. mRNA and protein expression of ACE, chymase, renin, AT1, and AT2 receptors were investigated by semi‐quantitative RT‐PCR and Western blot, respectively. Ang II secretion was examined by enzyme‐linked immunosorbent assay in condition medium.ResultsIH upregulated ACE (P=0.04) and chymase (P=0.05) expression while the expression of renin (P=0.94), AT1R (P=0.30), and AT2R (P=0.58) receptors did not change in HWP. Additionally, IH increased Ang II secretion in conditioned media (P=0.01).ConclusionsThis is the first study to demonstrate intermittent hypoxia‐mediated upregulation of Ang II generating enzymes ACE and chymase, along with increased AngII secretion in HWP. Together, our data strongly support the role of IH in upregulating RAS via increases in AngII. These studies suggest a role for adipose tissue and RAS in OSA related pathophysiology.Support or Funding InformationNIH R01 grant (#HL65176); Becari C. is supported by grant from CNPq‐Brazil (# 203802/2014‐4).
The Effects of Hyperglycemia on Insulin Resistance in a Drosophila model of Alzheimer's disease
Authors: Cline Delaena , Paddock Brie 
Published: November 2023

Alzheimer's disease (AD) and Type‐2‐Diabetes Myelitis (T2DM) are two increasingly prevalent diseases throughout the world, with evidence suggesting T2DM is a risk factor for AD. Alzheimer's disease, a neurodegenerative disorder, is characterized by the accumulation of A‐β leading to increased loss of memory and decreased synaptic plasticity. Hyperglycemia, associated with T2DM, leads to an exacerbation of amyloid accumulation and deposition within the brain. This deposition increases the likelihood of developing AD in these individuals and worsens present symptoms of those already suffering, including memory loss. It is unknown if hyperglycemia precedes amyloid‐induced pathogenesis or if accumulation of amyloid protein results in abnormal glucose handling. This study seeks to clarify this uncertainty by testing how high levels of A‐β within the nervous system of Drosophila melanogaster will affect the use of insulin in processing glucose. Using a standard Oral Glucose Tolerance Test, we are measuring the ability of this AD model to process a glucose challenge after 16 hours of starvation. Our current data indicates higher levels of circulating glucose in the AD model. These data suggest that coexpression of human amyloidogenic proteins in the nervous system may result in glucose handling abnormalities similar to those of T2DM.Support or Funding InformationCollege of Arts and Sciences, Arcadia University
Mitogen‐inducible Gene 6 Accelerates the Progression to Diabetes by Inhibiting EGFR‐Mediated Repair Mechanisms
Authors: Fong Kimberley M , Fueger Patrick T 
Published: November 2023

Type 1 diabetes (T1D) is caused by autoimmune‐mediated beta cell destruction. Following beta cell injury, the pancreas initiates a cellular repair or regeneration program, which includes epidermal growth factor receptor (EGFR) signaling. However, upon irreparable beta cell damage, EGFR signaling is dampened, disrupting attempts to restore functional beta cell mass and maintain normoglycemia. We have previously demonstrated that the negative feedback inhibitor of EGFR, Mitogen‐inducible gene 6 (Mig6), is induced by the pro‐inflammatory cytokines central to the autoimmune‐mediated beta cell destruction. We also established that pro‐inflammatory cytokines suppress EGFR activation, and siRNA‐mediated suppression of Mig6 restores EGFR signaling. Thus, we hypothesized that a loss of pancreatic Mig6 would protect mice from a chemically‐induced form of diabetes by enhancing EGFR signaling and promoting cellular repair. To this end, we treated mice lacking pancreatic Mig6 (PKO) and their wild‐type littermates (WT) with multiple low doses of streptozotocin (STZ) to induce beta cell death and diabetes or with saline as a control. Whereas STZ‐treated WT mice became hyperglycemic and had reduced beta cell mass, STZ‐treated PKO mice remained euglycemic and glucose tolerant, due to preserved beta cell mass. In addition, nitric oxide (NO) synthase inhibition blocks both the induction of Mig6 and cytokine‐impaired EGFR signaling, and treatment with an NO donor alone induces Mig6. Thus, we conclude that Mig6 ablation promotes beta cell damage repair, hence abating the progression to T1D. Our work suggests that Mig6 may be a novel therapeutic target for preserving beta cell mass in T1D.Support or Funding InformationThis work was supported by NIH grant DK099311 (PTF)
Regional Differences of Pancreatic Islet Area and Beta Cell Density in Obese and Lean Zucker Rats
Authors: Goudy Gabriel , Asano Shinichi , Hoskinson Hannah , Dawson Mariah , Breit Matthew , Olfert Mark , Frisbee Jefferson C , Chantler Paul D , Alway Stephan , Bryner Randall 
Published: November 2023

Several studies have characterized the density of beta‐cell mass in rodents reporting greater amounts in the body and tail (which is most proximal to the spleen) compared to the head of the pancreas (located in curved of the duodenum). Whether or not obesity alters the distributions within the pancreas is not known.PURPOSEThe objective of this study was to evaluate islet area and beta‐cell density in a rat model of obesity and metabolic syndrome.METHODSObese (OZR, n=8) and lean Zucker rats (LZR, n=8) were euthanized at 17‐weeks of age. Pancreases were harvested, sectioned into three regions (head, body, tail) and flash frozen in liquid N2 cooled isopentane. Frozen 7μm thick sections of pancreas tissues were cut, mounted on glass slides, and incubated with an insulin antibody that was labeled with Alexa‐Fluor 546 to identify the beta cells. The tissue was covered in DAPI to label the cell nuclei. Digital images of the pancreas cells were obtained by a Zeiss LSM 510 Confocal microscope using Zenn 2009 software. The insulin positive area of the stained islets was measured using Image J. The images of stained cells and islets were merged and the number of beta cells per islet recorded. ANOVA was used to compare the insulin positive area (islet area), total beta cell number, and beta cell density (beta cell/μm2 insulin positive area) between obese and lean rats with significance set at P<0.05.RESULTSThere was a significant main effect of phenotype (obese vs lean) for all three dependent variables (Table 1) with overall values lower in the OZR. There was also a significant interaction for phenotype and insulin positive area (islet area) as well as an interaction for phenotype and total beta cell number, but not for beta cell density. This suggests the biggest difference associated with obesity within the pancreas is in the tail region causing a reduced islet area.CONCLUSIONThese data indicate that obesity can significantly reduce islet area but that its affect is primarily in the tail region of the pancreas. The heterogeneity of islet area is consistent with previous findings among the different regions in the pancreas of the rat. Future research needs to examine what mechanisms are responsible for this regional difference.Support or Funding InformationNIHAHA
Mean±SE

Group (n=8)
Mean Islet Area (μm2)
Total Beta Cells
Beta cell/μm2 islet area

OZR
LZR
OZR
LZR
OZR
LZR

Head
6859 ±859
8266 ±266
82±26
115±15
0.0124 ± 0.0008
0.0144 ± 0.0008

Body
7831 ±831
7335 ±335
94±4
98±83
0.0123 ± 0.0010
0.0139 ± 0.0008

Tail
4749 ±651*
9475 ±475
56 ±6
136 ±3
0.0118 ± 0.0007
0.0147 ± 0.0004
Effect of dihydrotestosterone treatment on young and elderly mice skeletal muscle
Authors: Wendowski Oskar 
Published: November 2023

BackgroundSarcopenia, the progressive decline in skeletal muscle mass and function with age, is a debilitating condition. It leads to inactivity, increased susceptibility to falls and eventually to loss of independent living. Despite this, the causes and mechanism(s) underlying sarcopenia are still poorly understood.MethodsSmall muscle fibre bundles isolated from the extensor digitorum longus (edl; a fast‐twitch muscle) and the soleus (a slow‐twitch muscle) of young, old and elderly mice were used to investigated the effects of age and dihydrotestosterone (DHT) treatment on the expression and function of the sodium‐coupled neutral amino‐ amino acid transporter (SNAT) 2 and the sodium‐independent L‐type amino‐acid transporter (LAT) 2.ResultsAt all ages investigated, protein synthesis was ~2 times higher in slow‐twitch fibres than in the fast‐twitch fibres. Ageing led to a decrease in protein synthesis in both fibre types. However, the decline was greater in the fast‐twitch than in the slow‐twitch fibres. Moreover, it was accompanied by a reduction in the expression of both SNAT 2 and LAT2 proteins. The decrease in SNAT 2 protein was greater in the fast‐twitch fibres than in the slow‐twitch fibres. In contrast, ageing had no effect on the expression of SNAT2 and LAT2 mRNA. Neither did it affect the uptake of the non‐metabolisable amino‐acid, α‐methylamino‐isobutyric acid. Treating the muscle fibre bundles with physiological concentrations of DHT significantly increased protein synthesis in the fast‐twitch fibres of both young and elderly mice and that of the slow‐twitch fibres in elderly mice only. At both protein and mRNA levels, 5α‐reductase (SRD) 1 was more abundant in skeletal muscle than 5α‐reductase (SRD) 2. It was expressed mostly in skin, tibialis anterior and soleus muscle and its expression increased significantly with age. Chloroquine treatment abolished the increased protein synthesis caused by DHT treatment.ConclusionFrom these results we suggest that sarcopenia arises from an age‐dependent reduction in protein synthesis caused by lack of anabolic stimulus. We also suggest that muscles such as soleus and the tibialis anterior are resistance to sarcopenia because they are able to synthesis more DHT locally. The mechanism of action involves the recruitment of SNAT2 from an intracellular vesicle pool into functioning membrane transport proteins.
Drugs That Inhibit The Sarcoplasmic Reticulum Ca2+ ATPase (SERCA) and Prevention of Breast Cancer Cell Proliferation
Authors: Loulousis Melanie , Krager Stacey L , Darcy Yuanzhao L , Tischkau Shelley A , Copello Julio A 
Published: November 2023

Breast cancer is the most common cancer in women and a leading cause of death worldwide. Pharmacological modulation of intracellular Ca2+ homeostasis has been proposed as a potential target for therapeutic intervention for breast cancer. In this regard, thapsigargin and cyclopiazonic acid inhibit the Ca2+ ATPase of sarcoplasmic reticulum (SERCA) that pumps Ca2+ into the endoplasmic reticulum (ER) intracellular Ca2+ stores and display high efficacy for killing breast cancer cells. However, these “classic” SERCA blockers are highly toxic and also kill normal cells, thereby preventing use for anticancer therapy. We have determined that new drugs in the benzothiazepine (BZT) family (K201, CGP37157), as well as agents like pimozide and sipatrigine can inhibit SERCA through a novel Ca2+ dependent mechanism. These agents would not affect Ca2+ sequestration in ER when cytosolic Ca2+ levels are high, but potently inhibit this process when cytosolic Ca2+ approaches resting cell levels, which could help prevent ER overload under pathological conditions (ischemia) and provide cellular protection in normal tissues. Therefore, we tested the effectiveness of these drugs to inhibit growth of two breast cancer cell lines, MCF‐7 and MDA‐231, in culture. Our preliminary studies indicate that K201 inhibits MCF‐7 breast cancer cell growth and synergizes with anti‐cancer drugs to increase their effectiveness in killing these cells; thus, MCF‐7 cells are not “protected” by the decreased ER Ca2+ load provided by the SERCA inhibitors; in fact these drugs make them less viable. Building upon this data, we compared the effects of a “classic” SERCA blocker (thapsigargin) to the Ca2+ dependent SERCA blockers above on MCF‐7 and MDA‐231 breast cancer cells. We determined that all SERCA blockers effectively disrupted the metabolic activity of MCF‐7 cells but were much less effective in MDA‐231. In MCF‐7 cells, SERCA blockers also increased the effects of chemotherapeutic drugs (cisplatin, doxorubicin). The results indicate that targeting SERCA has potential to interfere with cancer cell growth. However, our results also suggest that resistance to chemotherapy in MDA‐231 may relate to these cancer cells becoming less dependent on intracellular Ca2+ signaling. We have also determined some of the structural characteristics that are required for BZT to block SERCA. Finally, we also found that drugs reported to do both, protection of normal cells in ischemia and inhibition of cancer cell proliferation, can also display significant SERCA blocking ability. These agents may serve as potential templates for developing new anti‐cancer drugs that target intracellular Ca2+ homeostasisSupport or Funding InformationAPS Frontiers in Physiology Research Fellowship to MLSIUSOM Team Development Award to SAT and JAC
Phosphorylation of α‐actinin‐4 (ACTN4) at serine 159 mimics the biochemical and cellular effects of human kidney disease mutations
Authors: Feng Di , Krishnan Ramaswamy , Stella Lorenzo , Schenone Monica , Hartigan Chrisina R , Pollak Martin R. 
Published: November 2023

α‐actinin‐4 gene (ACTN4) mutations cause a form of kidney injury called familial focal segmental glomerulosclerosis (FSGS) in humans. Individuals who carry disease‐causing ACTN4 mutations initially present with proteinuria due to podocyte injury, and many eventually develop end stage kidney disease. The disease‐causing ACTN4 mutations are exclusively located within the actin‐binding domain (ABD) of the encoded protein and result in increasing actin binding. We hypothesized that post‐translational modification of ACTN4 leads to similar biochemical and cellular changes observed in the disease‐causing ACTN4 mutant protein. Using mass spectrometry, we detected ACTN4 is phosphorylated at the serine 159 site (S159) in cultured human podocytes. By molecular dynamics simulations, we found that phosphorylated S159 causes a relative rotation of CH1 and CH2 domains within the ABD of ACTN4, potentially altering the actin binding interface. Supporting this is our observation that phosphomimetic S159D ACTN4 protein (which mimics the effect of phosphorylation at S159) increased F‐actin bundling activity in comparison with WT ACTN4, similar to the effect of kidney disease‐causing mutant K255E ACTN4 in vivo. Furthermore, when expressed in cultured human podocytes, phosphomimetic S159D ACTN4 led to similar changes as seen in the K255E mutant ACTN4: more prominent formation of actin stress fibers and significantly higher contractile force measured by traction force microscopy. We conclude that phosphorylation of ACTN4 at S159 induces characteristics similar to the kidney disease‐causing ACTN4 mutant both in vitro and in vivo. These data provide evidence that phosphorylation of ACTN4 plays an important role in regulating the cytoskeletal dynamics and function of podocytes.Support or Funding InformationR37DK059588 and T32DK007199
Integrating GWAS And Experimental Data To Predict Targets And Models For Mechanistic Validation Of Genetic Determinants Of Salt Sensitive Hypertension
Authors: Hultström Michael , McBride Martin W. 
Published: November 2023

The number of causal genes for human traits and diseases identified in GWAS increases steadily. However, only a small fraction has been successfully validated in experimental models to show how they may affect blood pressure regulation. Using recent hypertension GWAS we identified 1445 first and second tier SNPs with p‐values <10−5. Out of these 1379 could be annotated using SNP‐nexus and matched to 253 genes in Ingenuity Pathway Analysis (IPA).The first experimental dataset used were the Mouse Diversity Array SNP data from the Mouse Genome Database for C57BL/6J that has a salt sensitive blood pressure, and Balb/c that is much less salt sensitive. This identified 6253 SNPs, whereof 4548 were annotated through SNP‐nexus. In IPA the SNPs could be matched to 406 genes. Three genes with different alleles between the mouse strains were also found in the 253 genes from the GWAS dataset (HHAT, PARK2, ABCC2).Salt‐sensitive hypertension is usually thought to be associated to kidney function, and indeed all three gene‐products are detected by immunohistochemistry in renal tubuli in the proteinatlas.org database, where PARK2 also shows robust glomerular staining. Finally, in the results of an earlier micro array study we found that ABCC2 shows differential expression in the kidney between the strains, and is up regulated by a high‐sodium diet in C57BL/6J but not Balb/c indicating a functional difference.ABCC2 deficient mice are available on the C57BL/6 background but do not seem to have any baseline differences in blood pressure or sodium balance. However, they have mostly been used to study bile acid, and have not been studied under high‐ or low‐sodium conditions.In conclusion, ABCC2 is a second tier GWAS hit for hypertension in humans and appears amenable to mechanistic validation in the C57BL/6 mouse strain.Support or Funding InformationSwedish Society of Medical Research
Calcium‐sensing Receptor, Coupled with Transient Receptor Potential Canonical Channels 6, Regulates Cytosolic [Ca2+] in the Development and Progression of Pulmonary Hypertension
Authors: TANG HAIYANG , Yamamura Aya , Fraidenburg Dustin R , Song Shanshan , Yamamura Hisao , Chen Jiwang , Gu Yali , Ayon Ramon J , Black Stephan M , Garcia Joe G.N. , Makino Ayako , Yuan Jason X.‐J. 
Published: November 2023

RationaleElevated pulmonary arterial pressure (PAP) in patients with pulmonary arterial hypertension (PAH) is mainly caused by increased pulmonary vascular resistance (PVR), due primarily to sustained pulmonary vasoconstriction and excessive pulmonary vascular remodeling. An increase in cytosolic free Ca2+ concentration ([Ca2+]cyt) in pulmonary arterial smooth muscle cells (PASMCs) is not only a major trigger for pulmonary vasoconstriction but also an important stimulus for pulmonary vascular remodeling. Our previous data showed that the extracellular calcium sensing receptor (CaSR) expression and function were both enhanced in PASMC from idiopathic PAH patients. However, the implication of CaSR in the development of PAH remains elusive.MethodsCasr double knockout mice (Casr−/−Pth−/−) and transient receptor potential canonical Channels 6 (Trpc6−/−) have been exposed to hypoxia for 4 weeks to develop pulmonary hypertension. Following hypoxic exposure, right ventricle systolic pressures (RVSP), right ventricular hypertrophy by RV/(LV+S) ratios, and small pulmonary artery smooth muscle layer thickness were measured. The PAP was measured using the isolated perfused/ventilated mouse lung system. PASMCs were isolated from patients with IPAH, followed with [Ca2+]cyt measurement, proliferation and migration assays.ResultsIn the current study, we found blockade of TRPC6 channels significantly inhibits Ca2+‐induced increase in [Ca2+]cyt and OAG caused a larger increase in [Ca2+]cyt in IPAH PASMCs. Overexpression of CaSR and TRPC6 channels increase the extracellular Ca2+‐induced [Ca2+]cyt in normal PASMCs. Compared to wild mice, both Casr−/−Pth−/−and Trpc6−/− mice had significantly lower RVSP, less right ventricular hypertrophy, and displayed less pulmonary vascular remodeling after exposure to hypoxia. Deletion of Casr or Trpc6 gene significantly attenuates acute hypoxic vasoconstriction.ConclusionsThese data indicate that CaSR functionally interacts with TRPC6 and regulate [Ca2+]cyt in IPAH‐PASMC. Deletions of CaSR or TRPC6 prevent the development of hypoxia‐induced PAH in mice via its modulation of extracellular Ca2+ entry in PASMC, suggesting that CaSR may be a novel therapeutic candidate for the treatment of PAH.
Hypoxia Decreases Trx2 Protein and Activity in Pulmonary Vascular Cells
Authors: Wade Brandy E , Adesina Sherry E. , Hart C. Michael , Sutliff Roy L. 
Published: November 2023

Pulmonary Hypertension (PH) is characterized by a sustained increase in pulmonary arterial pressure. Increased pulmonary vascular pressure and resistance result in right ventricle (RV) hypertrophy and can ultimately lead to RV failure and death. The mechanisms leading to vascular cell proliferation and vascular remodeling in PH are incompletely defined. Current therapeutic approaches that stimulate vasodilation inadequately address the cellular proliferation and vascular remodeling that underlies the pathogenesis of PH. To more directly address mechanisms of cell proliferation in PH, this project focuses on the ubiquitin proteasome system (UPS) which plays an important role incellular homeostasis by regulating protein stability. Current evidence demonstrates changes in UPS activity in PH; however, the pathways and proteins influenced by these changes are poorly defined. Mice exposed to normoxic or hypoxic conditions for 3 weeks were used to identify proteins with altered ubiquitination in response to hypoxia exposure. Lungs were collected, and the PTMScan® Ubiquitin Remnant Motif (K‐ɛ‐GG) Kit was used to precipitate ubiquitinated proteins for analysis by mass spectrometry (MS). 245 proteins were identified with a ±2 fold change. In silico analysis identified several proteins that promote cell proliferation. Tax1bp1, Hspa8, Hspb1, Fhl1, Gnb1 interact with important cellular proliferation pathways and exhibit decreased ubiquitination in lungs from mice exposed to hypoxia, suggesting an increase in their stability in hypoxia. These results using MS and in silico analysis suggest that hypoxia‐induced changes in the UPS may impact proteins involved in pathways that regulate cellular proliferation by increasing or decreasing their stability.Support or Funding InformationThis work was supported by NIH RO1 HL 102167 and the T32 Training Program in Academic Pulmonary Medicine (T32HL076118).