bluuebunny/crossref_metadata_2025 huggingface dataset:
Behavioral effects of the novel benzodiazepine analog methyl 8‐ethynyl‐6‐(pyridin‐2‐yl)‐4H‐benzo[f]imidazo[1,5‐a][1,4]diazepine‐3‐carboxylate (MP‐III‐024)
Authors: Fischer Bradford , Hamade Bryan , Poe Michael , Cook James
Published: June 2021

MP‐III‐024 is a benzodiazepine (BZ) analog that, based on pharmacophore modeling, is predicted to exhibit agonist activity at the BZ site on the GABA‐A receptor. The present study assessed the behavioral effects of MP‐III‐024 on four endpoints in C57BL/6 mice. First, the antihyperalgesic effects of MP‐III‐024 were assessed in a model of inflammatory pain. Here, mechanical sensitivity was assessed following subcutaneous injection of the yeast extract zymosan A into the plantar surface of the footpad. MP‐III‐024 (1 – 32 mg/kg) produced dose‐ and time‐dependent reversal of mechanical sensitivity on this endpoint. Second, the anxiolytic‐like effects of MP‐III‐024 were assessed in a Vogel conflict procedure. In this procedure, MP‐III‐024 produced dose‐dependent increases in rates of suppressed responding up to a dose of 32 mg/kg, and decreases in responding below vehicle levels were observed at 100 mg/kg. Next, ambulatory behavior was assessed in locomotor activity chambers. In this procedure, MP‐III‐024 (1 – 32 mg/kg) failed to produce effects that were significantly different from vehicle. Finally, the response rate‐decreasing effects of MP‐III‐024 were examined in an assay of schedule controlled responding. Across the dose range tested, MP‐III‐024 (10‐100 mg/kg) failed to alter response rates in this procedure. Together, these observations will help provide a framework for studying GABA‐A receptor pharmacology which in turn should help guide the development of improved therapeutic agents for the treatment of pain‐ and anxiety‐related disorders.
Sunitinib Induces Growth Inhibition and Apoptosis in Breast Cancer MDA‐MB‐231 Cells through FOXO3a Signaling Pathway
Authors: Belali Osamah , Ansari Mushtaq , Korashy Hesham
Published: June 2021

Sunitinib (SUN) is a new multi‐targeted oral tyrosine kinase inhibitor that been recently approved against gastrointestinal stromal tumors and advanced renal cell carcinoma. Yet, the protective effect of SUN against breast cancer is poorly investigated. In this study, we have investigated the anti‐cancer properties of SUN against breast cancer and the possible role of Forkhead box type O (FOXO3a) transcription factor in SUN‐mediated effect using MDA‐MB‐231 cells as an in vitro model. Treatment of MDA‐MB‐231 cells with SUN caused a dose‐dependent cell growth suppression due to apoptosis as mediated by activation of caspase‐3 and p53 mRNA and protein levels using quantitative polymerase chain reaction and Western blot analyses. In addition, the percentage of cells undergoing apoptosis/necrosis determined by staining with annexinV–FITC and PI using flow cytometer was increased significantly. This inhibitory effect of SUN on MDA‐MB‐231 cell growth was associated with cell cycle arrest, downregulation of cell proliferator genes such as CyclinD1, and induction of oxidative stress markers such as heme oxygenase 1. Importantly, knockdown of FOXO3a expression using small interfering RNA (siRNA) significantly rescued MDA‐MB‐231 cells from SUN‐induced cell‐proliferative arrest, apoptosis, and oxidative stress at the mRNA and protein expression levels. In conclusion, the results clearly suggest that SUN‐induced inhibition of cell proliferation and apoptosis in MDA‐MB‐231 breast cancer cells is mediated through modulation of FOXO3a expression.
Evolution of Transcriptional Regulation of Duplicated Fatty Acid‐Binding Protein Genes by PPARs in Zebrafish: a Pharmacological Approach
Authors: Laprairie Robert , Denovan‐Wright Eileen , Wright Jonathan
Published: June 2021

Duplication of genes is thought to facilitate increasing organismal complexity. Duplicated genes may be retained in the genome if one of the duplicates acquires a novel function, or the functions of the ancestral gene are subdivided between the duplicates. A likely process for the retention of duplicates is the loss or gain of regulatory elements in the promoters of duplicated genes.

The objective of this study was to explore the evolution of gene promoters using the multigene family of fatty acid‐binding proteins (fabp) in zebrafish. Previous studies had implicated the peroxisome proliferator‐activated receptors (PPARs) in the regulation of some fabp genes. The promoters of the zebrafish fabp1a, fabp1b.1 and fabp1b.2 genes, duplicated by a whole genome duplication (fabp1a and ancestral fabp1b) followed by a tandem gene duplication (fabp1b.1 and fabp1b.2), were cloned into firefly luciferase reporter plasmids, transfected into HEK cells, and tested in the presence of PPARα‐ and PPARγ‐selective agonists. Expression of these genes was further analyzed by qRT‐PCR in explant cultures of zebrafish liver and intestine treated with PPAR agonists.

fabp1a expression was selectively increased by PPARα‐agonism in HEK cells and intestine. fabp1b.1 expression was selectively increased by PPARγ‐agonism in HEK cells and liver. fabp1b.2 promoter activity was not induced by PPAR modulation. The differential PPAR regulation of the duplicated fabp1 genes provide evidence of divergence of regulatory elements in these gene promoters, which may account for the retention of the duplicated fabp1 genes in the zebrafish genome.

Funding from NSERC, CIHR, and Dalhousie University.
Nanoformulations of Carfilzomib for Improved Metabolic Stability and Anti‐Cancer Efficacy
Authors: Ao Lin , Reichel Derek , Hu Di , Jeong Hyun‐Young , Kim Kyung Bo , Lee Wooin , Bae Younsoo
Published: June 2021

Carfilzomib (Cfz) is a second‐generation proteasome inhibitor drug approved for the treatment of multiple myeloma. In contrast to its promising anti‐myeloma activity, Cfz has shown limited efficacy in solid cancer patients, which may be in part due to its poor metabolic stability and short half‐life in vivo. Additionally, preclinical data suggest that prolonged administration of Cfz may be more effective than the current bolus dosing strategy. Thus, we hypothesized that Cfz nanoformulations with improved metabolic stability and sustained drug release profiles may enhance anti‐cancer efficacy. To test our hypothesis, we first prepared tethered nanoassembly (TNA) and micelle‐based formulations of Cfz and compared the rates of in vitro NADPH‐dependent Cfz metabolism in liver microsomes. Our results indicated that micelle‐based formulations were superior in improving metabolic stability to TNA formulations. Based on these results, we next prepared six micelle‐based formulations of varying polymer weight ratios and excipient modifications. All six micelle‐based formulations displayed improved Cfz metabolic stability and prolonged Cfz release profiles in vitro. Results from cell viability assays indicated that some of these nanoformulations can induce cancer cell death more effectively than free Cfz. In summary, we report that micelle‐based formulations of Cfz can achieve sustained drug release and improved metabolic stability. Taken together, these nanoformulations may have potential in expanding the therapeutic utility of Cfz.
In Vivo Evaluation of Two 5‐Aminosalicylic Acid Derivatives Considered as Potential Therapeutic Agents
Authors: Gutierrez Sanchez M , Rosales Hernandez M , Padilla Martinez I , Correa Basurto J , Mendieta Wejebe J
Published: June 2021

Several studies have shown that myeloperoxidase (MPO) is intimately involved in the initiation and progression of many inflammatory diseases [Lee et al., 2013].

Based on the foregoing, our research group has shown recently that two new derivatives of the 5‐aminosalicylic acid (5‐ASA), the 5‐{[(2E)‐3‐bromo‐3‐carboxyprop‐2‐enoyl]amino}‐2‐hydroxybenzoic acid (1) and 5‐[(4‐carboxybutanoyl)amino]‐2‐hydroxybenzoic acid (2), inhibit the catalytic activity of MPO and have antioxidant capacity in vitro. Therefore, this study aimed to evaluate the inhibitory effect of these compounds on catalytic activity of MPO in vivo.

Compounds 1 and 2 were synthesized and characterized chemically. The acute toxicity was determined in two rodent species [Guía 425, OECD, 2008]. The edema induction model was performed in the mouse ear by using 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) [Salazar et al., 2011]. The MPO activity was determined using the oxidation method of O‐dianisidine in presence of hydrogen peroxide [Quaiyoom et al., 2012].

Both compounds were obtained with acceptable yields (70%). These showed be safe for their use in rat and mouse (DL50˃1098 mg/kg). Furthermore their administration in ears of TPA‐treated mice, led to a significant decrease of catalytic activity of MPO in comparison with the positive control (indomethacin).

Therefore, these results suggest continue with the preclinical study of both compounds, since these could be used in future as potential therapeutic agents for the treatment of inflammatory diseases.
iNOS Contributes to Trichloroethene‐Mediated Autoimmunity
Authors: Wang Gangduo , Wang Jianling , Wakamiya Maki , Ansari G.A.Shakeel , Khan M Firoze
Published: June 2021

Reactive nitrogen species (RNS) have been implicated in the pathogenesis of autoimmune diseases (ADs). However, the potential mechanisms by which RNS contribute to the pathogenesis of ADs remain largely unexplored. Earlier studies from our laboratory in MRL+/+ mice suggested a role of RNS in trichloroethene (TCE)‐mediated autoimmunity. To further evaluate the contribution of nitrosative stress, especially iNOS, in TCE‐mediated autoimmunity, we conducted studies in iNOS‐null MRL+/+ mice. iNOS‐null MRL+/+ mice were obtained by backcrossing iNOS‐null mice to MRL +/+ mice. Female MRL+/+ and iNOS‐null MRL+/+ mice were treated with TCE (10 mmol/kg, i.p., in corn oil, every 4th day) for 6 weeks; their respective controls received corn oil only. TCE exposure in MRL+/+ mice led to significantly increased iNOS mRNA in livers, iNOS protein in livers and sera, and increased formation of nitrotyrosine (NT) in both sera and livers. These increases were associated with significant increases in serum anti‐double stranded DNA (anti‐dsDNA) and anti‐nuclear antibodies (ANA). Expectedly, iNOS levels and NT formation were negligible in both iNOS‐null controls and TCE‐treated iNOS‐null MRL+/+ mice. Even though TCE treatment still led to increases in serum anti‐dsDNA and ANA in iNOS‐null MRL+/+ mice compared to untreated controls, interestingly, the increases in serum anti‐dsDNA and ANA induced by TCE in the iNOS‐null MRL+/+ mice were significantly less pronounced compared to that in MRL+/+ mice. The observed differential autoimmune responses to TCE exposure in MRL+/+ vs. iNOS‐null MRL+/+ mice further support the contribution of iNOS and RNS in TCE‐mediated autoimmunity.
Effects of augmented O‐GlcNAcylation on activation and differentiation of macrophages
Authors: Zanotto Camila , Olivon Vania , Mestriner Fabíola , Alves Filho José , Carneiro Fernando , Tostes Rita
Published: June 2021

Macrophages (MØs) are classified as classically‐(M1) or alternatively‐(M2) activated, based on their exposure to different fate‐determining mediators. The post‐translational modification of proteins by O‐GlcNAcylation (O‐GlcNAc) is highly dynamic and modulates cell‐signaling processes. Acute increases in O‐GlcNAc levels reduce the release of pro‐inflammatory mediators and regulate inflammatory processes by decreasing e.g. NF‐kB activation. This study tested the hypothesis that increased O‐GlcNAc levels favor polarization of MØs to the anti‐inflammatory/M2 phenotype. Macrophages obtained from bone marrow of male BALB/c mice were incubated with vehicle, glucosamine, the O‐GlcNAcase inhibitors PugNAc (100 µM) and Thiamet‐G (TMG, 1µM), LPS + IFN‐g (1mg/ml + 200ng/ml) for M1 or interleukin‐4 (IL‐4, 50ng/ml) for M2 polarization. Expression of polarization markers (F4/80 and CD206) was assessed by flow cytometry. Cellular viability was not affected by these compounds, as measured by MTT reduction assay. Incubation of MØs with TMG, glucosamine and PugNAc produced a time‐dependent increase in O‐GlcNAc levels, determined by western blot. Incubation of undifferentiated MØs with TMG, as well as with LPS (1mg/ml), for 24h significantly increased IL‐1b release. In M1‐differentiated MØs, stimulation for 24h with TMG further increased IL‐1b and IL‐6 mRNA expression. In M2‐differentiated MØs, TMG did not change arginase I expression. These preliminary results suggest that increases in O‐GlcNAcylation in 24h contribute to a pro‐inflammatory phenotype in macrophages. Our studies suggest that the O‐GlcNAc pathway as a potential therapeutic target in diseases associated inflammatory responses.
Cofactor Dependent Cooperation Between Dimeric hSULT1B1 Subunits
Authors: Tibbs Zachary , Falany Charles
Published: June 2021

The cytosolic sulfotransferases (SULTs) catalyze the transformation of drugs and steroids into sulfate esters, generally “flagging” the molecules for excretion from the cell and body. Despite the variable substrate specificities between hSULT isoforms, all isoforms share a conserved specificity for the sulfonate donor, 3′‐phosphoadenosine 5′‐phosphosulfate (PAPS). Interactions involving the cofactor's 3′‐phosphate ensure the SULTs are not vulnerable to inhibition by other ATP analogues in the cell. However, with this gain in specificity, the SULTs sacrifice catalytic efficiency due to the resulting high affinity for the reaction byproduct, 3′, 5′‐diphosphoadenosine (PAP). We hypothesize the SULTs counteract susceptibility to PAP inhibition by communicating in a cooperative manner between dimer subunits, the result of which is a half‐site reaction mechanism. Molecular dynamic simulations (MDS) of hSULT1B1 dimers indicate the binding of PAP(S) to a single dimer subunit elicits stabilizing effects on its dimeric partner. Coupled with in vitro data showing two distinct PAPS binding affinities for the hSULT1B1 dimer, these data strongly suggest cofactor binding stimulates communication between dimer subunits, altering the cofactor binding affinity of the dimer partner. Further results indicate the presence of PAP actually promotes the binding of PAPS to the SULT dimer, triggering a structural shift that favors the release of PAP from the dimer subunit. Together, the data imply SULT half‐site reactivity results from an oscillating mechanism that promotes PAPS binding and PAP release across the SULT dimerization domain. This oscillatory cycle could be crucial in maintaining SULT catalytic efficiency under physiological PAPS concentrations, which vary throughout the body.
Elementary Ca2+ release events through IP3 receptors in the intact endothelium
Authors: Sonkusare Swapnil , Nelson Mark
Published: June 2021

Inositol triphosphate receptor (IP3R)‐mediated Ca2+ release from endoplasmic reticulum is one of the most important mechanisms to increase intracellular Ca2+ in all the cell types in the body. We have identified local IP3R‐mediated Ca2+ signals (“pulsars”) at endothelial projections to the smooth muscle (Ledoux et al, PNAS, 2008), which are involved in the control of vascular function. The objective of this study was to elucidate the elementary properties of the IP3R Ca2+ signals in the intact endothelium. The IP3R channel activity was optically recorded at 100 frames/second in slit‐open, resistance‐sized mesenteric arteries from mice using fluo‐8AM as a Ca2+ indicator and EGTA‐AM to limit the spread of Ca2+. Step‐wise (quantal) changes in fluorescence were measured, which likely correspond to Ca2+ flux through a single IP3R channel. Comparison of quantal levels for the voltage‐dependent Ca2+ channel, IP3Rs and TRPV4 channel indicates an approximate single channel conductances to Ca2+ of 3.5 pS, 14 pS and 35 pS respectively. The muscarinic receptor agonist carbachol (CCh, 10 mM) increased the activity of IP3R events, but did not alter the quantal amplitude. The IP3R events were eliminated by cyclopiazonic acid (SERCA inhibitor) and U73122 (phospholipase C inhibitor). Our results indicate that a IP3R‐mediated Ca2+ pulsar reflects the opening of about 15 IP3R channels, and the presence of EGTA decreases this to about 7. These results may provide a unique method for studying the modulation of elementary properties of IP3Rs by physiological modulators in health and disease.
Gravin Scaffolding Protein Mediates Signaling Involved in theRegulation of Hyperlipidemia and Atherosclerosis
Authors: Fan Qiying , Wijaya Cori , Diaz Diaz Andrea , Vo Hiep , McConnell Bradley
Published: June 2021

Atherosclerosis is a leading health problem and contributes to cardiovascular related mortality. Furthermore, hyperlipidemia is a major atherogenic factor.Understanding the cellular mechanisms leading to atherosclerosis is still lacking. Gravin (AKAP12), an A‐kinase anchoring protein (AKAP), scaffolds and targets various signaling proteins to specific intracellular locations and has been implicated in mediating lipid metabolism. Thus, the purpose of this study was to determine whether the absence of gravin‐mediated signaling would prevent the initiation and progression of atherosclerosis.5 week‐old wild‐type (WT) and gravin‐truncated (gravin‐t/t) mice were subjected to high fat diet (HFD) or normal diet (ND) for 16‐weeks. Cholesterol, triglyceride and VLDL level in serum were significantly decreased in gravin‐t/t HFD compared to WT HFD mice. In addition, systolic and diastolic blood pressure was also lower in gravin‐t/t HFD mice.Furthermore, gravin‐t/t HFD mice showed lower liver‐to‐body weight ratio as well as decreased lipid accumulation and liver damage.These observations are consistent with the lower serum alanine aminotransferase(ALT) and aspartate aminotransferase (AST) levels identified in gravin‐t/t HFD mice. In addition, gravin‐t/t HFD mice had decreased expression of the active form of sterol‐regulatory‐element‐binding protein 2 (SREBP2), which resulted in lower HMG‐CoA reductase (HMGCR) and low‐density lipoprotein receptor (LDLR) expression. Finally, we observed less aortic lipid accumulation in gravin‐t/t HFD mice. Our data indicates that the absence of gravin mediated signaling were resistance to hyperlipidemia and prevented the progression of atherosclerosis.
Alterations in Uromodulin and Microtubule Associated Protein (MAP)7 in Human Renal Cell Carcinoma (RCC)
Authors: Banes‐Berceli Amy , Polina Aws , Singhal Sara , Jammerino Jennifer , Fulton Matthew , Geddes Timothy , Thibodeau Bryan , Wison George , Hafron Jason
Published: June 2021

There is urgent clinical need for better treatment options as RCC responds poorly to chemotherapy and currently has no early detection biomarkers available. We previously demonstrated increased activity of Janus Kinase (JAK2) pathway.We hypothesized that increased JAK2 activity resulted in increased levels of MAP7, linked to chemoresistance, and decreased levels of Uromodulin, a secreted protein linked to renal protection, measurable in the urine. We utilized samples from normal (n=14), benign (n=6), and histological Fuhrman grades: Grade 1 (n=3), Grade 2 (n=10), Grade 3 (n=14), and Grade 4 (n=3). We analyzed samples with Affymetrix human gene microarrays and Western Blot technology. The samples were all clear cell renal carcinomas verified by a pathologist. Altered gene and protein expression was detected by ANOVA taking into account histological stage. We found MAP7 mRNA to be highly expressed in the RCC samples compared to the controls and benign and it increased with severity of the cancer stage. Uromodulin mRNA levels were increased in the benign samples compared to control and were undetectable in all stages of the RCC samples. We saw corresponding changes in the protein levels of both MAP7 and Uromodulin. We also performed an ELISA to measure Uromodulin in the urine and found a significant decreases in stage 3 and 4 RCC samples compared to normal and benign. There were significant increases in Uromodulin in benign, stage 1 and 2 RCC samples compared to control. These data suggest that altered function of these protein may be involved in RCC. Zafarna Fund
Global Intracoronary Allogeneic Cardiosphere‐Derived Cells Promote Functional Repair Without Affecting Myocardial Perfusion in a Porcine Model of Acute Myocardial Infarction
Authors: Weil Brian , Goelz Andrew , Suzuki Gen , Palka Beth , Canty John
Published: June 2021

Objective

Intracoronary cardiosphere‐derived cells (icCDCs) exert beneficial effects on left ventricular (LV) function in experimental models of myocardial infarction (MI), but the role of microvascular remodeling and improvements in myocardial perfusion are unclear. We determined whether global icCDCs administered after reperfusion promote functional repair by improving myocardial blood flow in a swine model of acute MI.

Methods

Immunosuppressed swine (cyclosporine 100 mg/day, n=15) underwent a 60‐minute LAD occlusion. After 30 minutes of reperfusion, animals were randomized to receive vehicle or 20 x 106 allogeneic CDCs infused into the 3 major coronary arteries at 106 cells/min under continuous flow. LV function (CT) and myocardial perfusion (microspheres) were assessed at baseline and 1‐month after MI, at which point the heart was excised for histological assessment of microvascular density.

Results

Segmental wall thickening in the infarct, border, and remote myocardium was higher in icCDC‐ vs. vehicle‐treated animals (Table). As a result, there was a significant improvement in LV ejection fraction (57.7 ± 2.7 vs. 50.7 ± 4.1%, p<0.05). However, resting and adenosine‐dilated blood flow remained similar between groups, with a comparable density of arterioles and capillaries throughout the LV in each group.

image

Conclusion

The functional improvement afforded by allogeneic icCDCs administered immediately after acute MI is independent of angiogenesis with no changes in microvascular density or myocardial perfusion.

Support

National Heart Lung and Blood Institute (HL‐055324, 1F32HL‐114335)
Least Significant Change of Body Composition and Bone Mineral Density Measured by Dual energy X‐Ray Absorptiometry
Authors: Najm Nadia , Popp Collin , Traylor Daniel , Jesch Elliot
Published: June 2021

Introduction

This study is designed to determine the least significant (LSC), or the least amount of change that can be considered significant when measuring body composition with Dual Energy X‐ray Absorptiometry (DXA). In the past five years, researchers have focused on measuring the LSC of bone mineral DXA scans or body composition DXA scans in athletic populations. However, none have measured LSC for body composition in non‐ athletic populations or the LSC of each component of body composition.

Methods

Forty‐six adults (39 female; 7 male) partook in the study aged between 50.78 ± 15.27 years. They weighed 157.90 ± 38.83 pounds, and had a of height 64.83 ± 3 inches. Any participant who that was pregnant was excluded from the study. DXA scans for body composition and BMD was taken by the same technician. Each scan was consecutively taken 3 times, and after each scan the participant got up from the table, and was repositioned for the next scan. Results were calculated with the International Society of Clinical Densitometry's calculator.

Results

DXA scans for WB‐ Bone Area, WB‐ BMD, WB‐ Lean Mass + BMC, WB‐ Lean Mass, and WB‐ Total Mass measured less than 1% of coefficient variation, where as AP Spine BMD, WB‐ Bone Mass, WB‐Fat Mass, and WB‐% Body Fat were just over 1%, between 1.07‐ 1.3% coefficient variation. However, the WB‐ VAT Mass had a significantly higher percent (15.9%) coefficient of variation.

Conclusions

DXA scans for whole body and spine all come with a 1% error as reported by DXA manufacturers, with the exception of WB VAT mass that had a significantly higher percent error.
Homocysteine Elicits an Inflammatory Profile in Murine Macrophages Through an EMMPRIN Mediated Pathway
Authors: Winchester Lee , Veeranki Sudhakar , Givvimani Srikanth , Tyagi Suresh
Published: June 2021

Introduction

Macrophages are known to differentiate between two distinct phenotypic subsets: M1 and M2. The M1 phenotype elicits a pro‐inflammatory profile while M2 macrophages are categorized as being anti‐inflammatory. Hyperhomocysteinemia (HHcy) is associated with many inflammatory diseases such as obesity and atherosclerosis. It is known that HHcy increases specific macrophage secretory factors, such as MCP‐1 and MMP‐9. However, the direct effect of HHcy on macrophage phenotype is largely unexplored. We hypothesize that HHcy causes macrophages to shift into a more inflammatory phenotype or M1.

Methods

We utilized 2 murine macrophage cell lines: Raw 264.7 and J774A.1 cells. These cells were treated with 500 and 100 µM homocysteine, respectively, for 24 hours. Methods employed are Western Blot, ICC, FACS and PCR.

Results

In Raw 264.7 cells, 24 hour treatment with 500 μM Hcy resulted in a 2.25 fold increase in CD40 protein expression: a cell surface marker on M1 macrophages. ICC confirms this increase in CD40. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) was also increased by 1.62 fold, indicating that MMP‐9 induction with Hcy may be through an EMMPRIN mediated pathway. J774A.1 cells were more responsive to Hcy treatment, demonstrating a 1.4 fold induction of CD40 with 24 hours of 100 μM treatment. ICC demonstrates increased MMP9 production in this cell line. We suggest that Hcy may alter IL‐1β, iNOS, TIMPs 1 and 4, and MMP‐2, as well as EMMPRIN in MMP9 production.

Conclusions

Our current data suggests that Hcy treatment shifts macrophage phenotype towards the pro‐inflammatory, M1 subset. We believe that HHcy increases IL‐1 β, iNOS and MMP‐2, while demonstrating decreases in TIMPs 1 and 4.
Samchuleum suppressed high glucose‐induced renal proliferation and fibrosis in cultured rat mesangial cells
Authors: Yoon Jung Joo , Lee Yun Jung , Choi Eun Sik , Kim Mi Gyeong , Han Byung Hyuk , Kang Dae Gill , Lee Ho Sub
Published: June 2021

Diabetic nephropathy associated with morbidity and mortality of diabetic patients. Mesangial cell proliferation is known as the major pathologic features such as glomerulosclerosis and renal fibrosis. Samchuleum (SCE), originally recorded in an ancient Korea medicinal book named “Donguibogam”, is a well‐known blended traditional herbal formula. Thus, this study was investigated the inhibitory effect of SCE (1‐50 μg/ml) on high glucose (HG)‐stimulated rat mesangial cells proliferation and fibrosis.

Thymidine incorporation under HG was significantly accelerated, which was inhibited by SCE in a dose dependent manner. Pre‐treatment of SCE induced down‐regulation of cyclins/CDKs and up‐regulation of CDK inhibitor, p21waf1/cip1 and p27kip1 expression. In addition, SCE significantly suppressed the HG‐induced ROS production. Thus, SCE consequently inhibited HG‐induced mesangial cell proliferation through the inhibition of ROS signaling pathway.

HG enhanced expression of ﬁbrosis biomarkers such as collagen IV and CTGF, which was markedly attenuated by SCE. Moreover, SCE inhibits HG‐induced fibronectin mRNA expression. SCE decreased TGF‐β1 and p‐Smad2/Smad4 expression, whereas increased Smad7 expression under HG. Thus, SCE promoted ECM degradation through disturbing TGF‐β–SMAD signaling.

In conclusion, these results suggested that SCE has a protective effect on renal proliferation and fibrosis. Thus SCE might prove to be effective in the treatment of renal dysfunction leading to Diabetic nephropathy.
Cardiac Mechanoreceptor Activation in Humans: Evaluation by Phenylephrine and Norepinephrine
Authors: Sørensen Henrik , Ogoh Shigehiko , Sato Kohei , Siebenmann Christoph , Rasmussen Peter , Lundby Carsten , Secher Niels
Published: June 2021

Aortic occlusion activates cardiac mechanoreceptors that elicits muscle vasodilatation and reduce heart rate (HR). Norepinephrine and phenylephrine reduce HR but whether muscle blood flow increases is not known.

Cardiovascular variables were determined in males during administration of norepinephrine (n=15) or phenylephrine (n=7) and during 50° head up tilt (HUT) with and without phenylephrine (n=7). Mean arterial pressure (MAP), stroke volume (SV), HR, and total peripheral resistance (TPR) were derived by Modelflow® and near infrared spectroscopy determined biceps muscle oxygenation (SmO2).

With norepinephrine MAP, SV, and TPR increased by 24 mmHg (18‐31; P<0.001; median; interquartile range), 13 ml (0‐22; P=0.026), and 3 mmHg min‐1 l‐1 (1‐4; P=0.004). HR was reduced 4 bpm (2‐7: P=0.025), while SmO2 increased 7% (3‐10; P=0.002). Similarly with phenylephrine MAP, SV, and TPR increased by 31 mmHg (22‐43; P=0.016), 18 ml (13‐24: P=0.031), and 7 mmHg min‐1 l‐1 (5‐9; P=0.016). HR was reduced by 19 bpm (17‐23; P=0.016) and SmO2 increased 7% (5‐9; P=0.016). During HUT SmO2 and SV decreased 14% (9‐14; P=0.016) and 19 ml (9‐23; P=0.031), HR increased (14 bpm; 3‐22; P=0.016), and TPR was stable. Yet, HUT phenylephrine reduces HR (15 bpm; 11‐19; P=0.016) as TPR increased (6 mmHg min‐1 l‐1 (4‐6; P=0.004) while SmO2 and SV were restored.

HR decreases and TPR increases with administration of norepinephrine or phenylephrine independent of cardiac preload. Nevertheless, SmO2 increases indicating that the reduction in HR is mediated by cardiac rather than arterial mechanoreceptors when stroke volume becomes elevated.
Glia Inhibitor Blocks Ventilatory Acclimatization to Chronic Sustained Hypoxia
Authors: Stokes Jennifer , Fu Zhenxing , Powell Frank
Published: June 2021

Neuroglial communication in the central nervous system (CNS) is proving to be an important component of neuronal signaling both in normal conditions as well as stressed or injured states. We investigated the role of CNS glial cells in neural control of ventilation during chronic hypoxia. To assess this, rats received daily administration of either minocycline, a selective microglia inhibitor, (45mg/kg) or saline (i.p.) beginning one day before and during 7 days of exposure to hypoxia (PO2 = 80 Torr). Normoxic controls, which received daily injections of either minocycline (45mg/kg) or saline, were not exposed to hypoxic conditions. Ventilation was recorded using barometric pressure plethysmography in unrestrained rats both before and after exposure to hypoxia. Daily administration of minocycline significantly (p < 0.01) decreased ventilation in normoxia and the hypoxic ventilatory response but not the hypercapnic ventilatory response, following exposure to chronic hypoxic conditions. Breathing frequency and ventilation were significantly lower in chronically hypoxic rats treated with minocycline compared to saline. Minocycline did not have an effect in normoxic conditions. Activation of microglia and astrocytes is being quantified in the NTS using immunohistochemistry. These results suggest a role for glial signaling in the physiological acclimatization during hypoxia.

Supported by NIH RO1 HL‐081823
The Natriuretic Response to Acute K+ Intake
Authors: Jensen Iben , Larsen Casper , Leipziger Jens , Sørensen Mads
Published: June 2021

Introduction

Diets rich in K+ are associated with unexplained protection against hypertension. Acute K+ intake induces natriuresis and kaliuresis in mammals linked to marked dephosphorylation of renal Na+/Cl‐‐cotransporters (NCC). Reduced activity of NCC increases Na+ delivery to the collecting ducts, hypothesized to increase driving force for ENaC‐dependent K+ secretion. Here we investigated K+ and Na+ excretion in mice adapted either to preserve or promote Na+ loss.

Methods

Mice were fed low (0,03%), control (0,2 % ) or high (2%) Na+ diet for 25 days. Weekly, mice received gavage of either K+ or vehicle. Mice were placed in metabolic cages and urine was collected real‐time. ENaC‐dependence of kaliuresis was assessed by benzamil injections prior to gavage.

Results: 1

Aldosterone and cleavage‐products of ENaC were inversely related to dietary Na+ content. 2 K+ excretion rate was reduced in mice on high Na+ diet compared to the other groups. 3 In all dietary groups, K+ load induced natiuresis and complete NCC dephosphorylation. However, maximal Na+ excretion rate was reduced 80% in Na+‐restricted and increased 15% in Na+‐loaded mice. 4 Benzamil prior to K+ loading increased natriuresis and decreased kaliuresis. Importantly, benzamil abolished all differences in kaliuresis and natriuresis between the groups.

Conclusion

Our data suggest that acute K+‐induced kaliuresis is completely ENaC‐dependent. Maximal K+ excretion rates are attenuated when ENaC is pharmacologically blocked or physiologically down regulated. To supply adequate luminal Na+ to the connecting tubules and the collecting ducts NCC is dephosphorylated following acute K+ load under all dietary Na+ regimes. This leads to natriuresis, even in severely Na+ restricted animals.
Hydrogen Peroxide Evokes Acute Calcium Influx in the Podocytes of Murine Glomeruli
Authors: Ilatovskaya Daria , Palygin Oleg , Staruschenko Alexander
Published: June 2021

Podocytes are key components of the renal filtration barrier: podocyte depletion is associated with progression of proteinuric diseases. The physiological function of these cells is critically dependent on proper intracellular calcium handling – excessive calcium influx may result in apoptosis and glomeruli damage. Podocyte loss is a primary characteristic for glomerulosclerosis, a condition common for diabetic nephropathy and other kidney diseases, which have been associated with elevated levels of reactive oxygen species (such as H2O2) in the kidney. The goal of this study was to investigate how changes in extracellular H2O2 affect podocyte calcium handling; in order to test this, we performed ex vivo confocal calcium imaging in the podocytes of the freshly isolated mouse glomeruli loaded with Fluo4 and FuraRed dyes. The basal level of calcium within the podocytes was found to be 145.2 ± 10.1 nM. A dose‐dependent increase in intracellular calcium was observed in response to application of H2O2 (2.5 µM to 300 µM), with the EC50 for the peak response being 56 µM. Interestingly, lower concentrations of H2O2 caused a transient with a decay within a 60 s range, whereas doses of 50 µM and higher evoked a stable plateau elevation in intracellular calcium level, which could be attributed to irreversible apoptotic effects. We further recruited patch‐clamp electrophysiology to measure the activity of calcium channels in this preparation. The open probability of the TRPC channels was acutely increased in response to application of 25 µM H2O2. Taken together, these data indicate that H2O2 evokes calcium influx in the podocytes through the ion channels of the TRPC family, and this mechanism might play an important role in the development of kidney damage.
Participation of Immune Cells in Klotho Deficiency‐induced Salt‐sensitive Hypertension
Authors: Zhou Xiaoli , Lei Han , Sun Zhongjie
Published: June 2021

Objective

Klotho (KL) is an aging‐suppressor gene. The purpose of this study is to investigate if immune cells are involved in KL deficiency‐induced salt‐sensitive hypertension.

Methods & Results

Haploinsufficiency of KL (+/‐) caused spontaneous and persistent increase in blood pressure (BP). Interestingly, the high salt (HS) intake further increased BP and exacerbated hypertension in KL(+/‐) mice, but not in WT mice, suggesting that KL deficiency elicited salt‐sensitive hypertension. The HS loading remarkably increased expression of MCP‐1 and infiltration of macrophages and T cells in kidneys in KL(+/‐) mice. Blockade of CC chemokine receptor 2 (CCR2) by a CCR2‐specific antagonist (INCB3284) abolished the HS‐induced increase in BP in KL(+/‐) mice, suggesting an important role of monocyte chemotaxis in salt‐sensitive hypertension. INCB3284 abolished infiltration of macrophages and T cells in kidneys in KL(+/‐) mice. The expression of Sgk1, NCC and ATP synthase β in kidneys was enhanced by HS loading in KL(+/‐) mice, which can be eliminated by INCB.  Blockade of CCR2 abolished HS‐induced renal structural damage and functional impairment in KL(+/‐) mice.

Conclusion

KL deficiency caused salt‐sensitive hypertension and renal damage via MCP‐1/CCR2‐mediated inflammation and infiltration of macrophages and T cells.
Regulation of Myelination in the Prefrontal Cortex by the Gut Microbiota: Implications for Health and Disease
Authors: Hoban Alan , Stilling Roman , Desbonnet Lieve , Shanahan Fergus , Dinan Timothy , Cryan John , Clarke Gerard
Published: June 2021

Background

Multiple lines of evidence suggest a potential role for the gut microbiome in multiple sclerosis, a degenerative demyelinating disorder. For example, germ‐free mice, which are microbiota deficient, appear to be protected against the development of experimental allergic encephalomyelitis (EAE). The gut microbiota can regulate gene expression at the level of the CNS. In this study, we aimed to establish whether germ‐free mice have alterations in genes of relevance to CNS myelination.

Methods

Using qRT‐PCR, we evaluated expression levels of myelin basic protein (Mbp), myelin oligodendrocyte protein (Mog), myelin oligodendrocyte basic protein (Mobp), proteolipid protein 1 (Plp1) and myelin associated glycoprotein (Mag) in relevant brain regions in conventionally raised, germ‐free and germ‐free colonised mice.

Results

Germ‐free mice displayed increased expression of all 5 myelinating genes only within the prefrontal cortex. Colonisation of the germ‐free animals immediately post‐weaning normalised the expression of these myelin structural protein genes. Germ‐free mice also displayed a 2 fold increase in expression of Egr2/Krox20, a transcription factor within the PFC known to be involved in mechanisms regulating myelin formation.

Conclusion

The gut microbiota can regulate myelin structural protein gene expression in the CNS. Moreover, our results suggest that the microbiota can be successfully targeted later in life to modulate myelination patterns. This raises the possibility that targeting the gut microbiota during critical time windows could be a viable approach to promote remyelination or to prevent further demyelination in disorders such as multiple sclerosis.
Attenuating heat stroke syndromes in rats by heat shock protein 72‐mediated hypobaric hypoxia preconditioning
Authors: Chang Ching‐Ping , Chang Ming‐Wen , Niu Ko‐Chi
Published: June 2021

Herein, we investigated whether hypobaric hypoxia preconditioning (HHP) improved the multiple organ dysfunction (MODS) in heatstroke by up‐regulation of heat shock protein 72 (HSP72). Anesthetized rats were randomly assigned to (a) non‐HHP+non‐heated group, (b) non‐HHP+heated group, (c) HHP+heated group and, (d) HHP+HSP72 antibodies(Abs)+heated groups. All heated groups were exposed to heat stress (43oC, 70 min) to induce heatstroke. For HHP, animals were exposed to 0.66 atmosphere absolute (18.3% O2) for 5 h daily for consecutive 5 days per week for 2 weeks before the onset of heat exposure. HHP significantly attenuated the MODS that occurred during heat exposure. In addition, the beneficial effects of HHP could be significantly reduced by HSP72 Abs preconditioning. Our results show that HHP protects rats from heat‐induced MODS via up‐regulating HSP72. The HHP could be a novel strategy for the prevention of heatstroke animals od patients before heat exposure.
Mitosis in Secretory Ameloblasts
Authors: Hogg Russell , Smith Timothy , Sanchez Adriana , Bly Karlie , Gografe Sylvia
Published: June 2021

Our objective is to test the dictum that secretory ameloblasts cannot divide. Dental biology textbooks often present this idea without citation or evidence; its source seems to be studies of rodent incisors from the 1960's. However, rodent incisors are ever‐growing, and not acceptable models for enamel epithelia in other mammals. There are two reasons to expect that ameloblasts divide. First, recent studies show that tensile forces induce mitosis in epithelia, and enamel epithelia likely undergo tension as enamel is deposited and the surface area of the growing crown expands. Second, we now know that ameloblasts are subject to the same biorhythms that synchronize proliferation of osteoblasts. If ameloblast division is also coordinated by the brain and occurs at night, it is unlikely to have been observed since most experimental protocols occur in daylight.

To test the hypothesis, we first measured enamel area from surface and enamel‐dentine junction in Saimiri sciureus tooth sections. We also measured enamel rod width at both sites, as a proxy for ameloblast width. We found an average 9% greater area at the tooth surface, but no difference in enamel prism width. We also searched for mitotic figures in tooth buds of perinatal Saguinus spp. with random times of death, but found none. In conclusion, increasing tooth surface area from deep to superficial and no change in cell size suggests that ameloblast mitosis is likely. However, we did not find any mitotic figures, possibly because of sample size and lack of control over time of death.
Osteocyte Morphology in the Primate Craniofacial Skeleton
Authors: Smith Leslie , Ren Yinshi , Feng Jerry , Dechow Paul
Published: June 2021

Frost's “mechanostat” theory suggests MES varies throughout the skeleton in order to maintain strength in low strain regions such as the cranial vault. More spherical osteocytes have been shown to produce more nitric oxide than oblong osteocytes undergoing the same stress in vitro. In this study we look at osteocyte shape differences in a high and low strain region of the human craniofacial skeleton: the zygoma (Z) and the browridge (BR), respectively, and between the BRs of a chimp and human. We hypothesize that BR osteocytes will be more spherical in shape than those in the Z, and that osteocytes in the robust chimp BR will be more spherical than those in the human. Rectangular samples were extracted from the BR and Z regions of a 36 year old human female and an adult female chimp BR. Samples were stained with FITC, embedded in MMA and captured using confocal microscopy. Osteocytes from each cortical region were analyzed in Imaris for differences in sphericity. T‐tests were used to determine significance. Osteocytes in all three regions were more spherical (average sphericity greater than 0.63) and less ellipsoidal. There were no significant differences found between either group. These results suggest that differences between postcranial and calvarial osteocytes that are useful in maintaining bone mass in regions of low strain may not exist between subregions of craniofacial bone.

Funding: National Science Foundation Physical Anthropology HOMINID program (NSF BCS 0725141)
Morphological Variation in the Communicating Branch between the Lateral and Medial Plantar Nerves
Authors: Tilden Sarah , Gravenstreter Jonathan , Michalak Samantha , Bee Mary
Published: June 2021

Studies investigating the presence of a communicating branch between the lateral and medial plantar nerves (LMCB) have found widely differing occurrence rates. In our study on human cadavers of Caucasian descent (ages 50‐103), feet were dissected to ascertain the presence, dimensions, and course trajectory of the LMCB. The LMCB was present in 77.3% of the feet, with most coursing in the medial oblique direction (58.8% of anastomoses). The medial oblique coursings started at a proximal location on the lateral plantar nerve, lateral to quadratus plantae, and ended distally on a branch of the medial plantar nerve between flexor digitorum brevis tendons to digits 3 and 4. Transverse and lateral oblique coursings were also found, constituting 17.7% and 23.5% of anastomoses each. The average length, width, and thickness of the LMCB were 11.6 mm, 1.1 mm, and 0.5 mm respectively. A single instance of two parallel running branches was also found, both coursing in the lateral oblique direction 14.0 mm apart, as was a branch that bifurcated laterally before anastomosing. A single branch originating from the deep branch of the lateral plantar nerve superior to quadratus plantae was also found. These results suggest a more frequent occurrence and a more widely varying morphology than has been previously described. The prevalence and position of this anastomotic nerve is important in planning podiatric surgical approaches, and invaluable in characterizing morphological variations in peripheral nerve coursings, networks, and anastomoses.
Thymoquinone Inhibits Neurodegeneration and Enhances Neurogenesis in Kainic Acid‐induced Middle‐aged Wistar Rat Model of Temporal Lobe Epilepsy
Authors: Brito Sijam , Kilarkaje Narayana , Rao Muddanna
Published: June 2021

Inflammation in brain is a causative factor for neural excitability and reduced neurogenesis in chronic epilepsy. Thymoquinone (TQ), an active constituent of Nigella sativa seeds, possesses anti‐inflammatory and neuroprotective roles. This study investigated the role of TQ (10 mg/kg; ip) in neuroprotection and neurogenesis in kainic acid (0.5µg; intraventricular)‐induced model of chronic temporal lobe epilepsy in middle aged Wistar rats (12‐monh‐old). The normal and sham controls, epileptic and TQ‐treated epileptic rats (n=6/group) were sacrificed at 5th day and at the end of 3rd month. Brain sections were stained with Fluro‐Jade B and labeled for Doublecortin. The number of degenerating neurons in CA1 (Cornu ammonis 1), CA3 and dentate hilus were quantified at 5th day, and newly born neurons and mossy fiber sprouting were quantified in the dentate gyrus at the end of 3rd month. TQ significantly reduced the neuron degeneration (25%), in CA1, CA3 and dentate hilus, suppressed the mossy fiber sprouting (30‐40%), and increased (20‐30%) the neurogenesis in dentate gyrus as compared to epilepsy‐only group (P<0.001). Our results suggest that TQ inhibits neurodegeneration and enhances neurogenesis in hippocampus and has anti‐epileptic effects (Supported by Kuwait University Grant #YM02/14).​
Vestibular and Proprioception Deficits in Adults with Controlled Diabetes Mellitus II Causes Postural Control Instability
Authors: Rosario Martin , Dunbar Donald , Ortiz Alexis
Published: June 2021

Objective

individuals with controlled type two diabetes (cDM) often are unaware of balance impairments, although these changes are occurring and may become impairments. Thus, the objective of this study is to make an individual comparison and characterization of each system involved during standing balance (vision (Vi), proprioceptive (Pro) and vestibular (Ve) tasks in people with DM and healthy‐control individuals and identify the most affected system.

Methods

Fourteen participants with DM (age 59 ± 5, BMI 29 ± 3 kg/m2and 8 ± 6 years since diagnosis) and eighteen healthy participants (age 57 ± 5, BMI 27 ± 3 kg/m2) participated in this study. Postural stability during still standing for 20 seconds was assessed using a pressure mat by measuring center of pressure (CoP). Static postural control was evaluated during eight sensory conditions that perturbed or stimulated the visual (Vi), proprioceptive (Pro) and vestibular (Ve) systems.

Results

A Mann‐Whitney U Test was used to compare CoP data between DM and normal subjects. When pitching the head with the eyes open while standing on the foam pad, CoP displacement was significantly greater in DM subjects, indicating increased body sway and less balance control (CoP displacement: 39.65 ± 18.6 cm2 in DM vs. 24.33 ± 9.5 cm2 in controls, P= 0.009).

Conclusions

The conditions in which the Ve and Pro systems were challenged, the group with DM had a greater CoP displacement when compared to the control group. This suggests that in patients with DM without a diagnosis of neuropathy, postural control changes are already detected making them unstable by affecting their balance.