bluuebunny/crossref_metadata_2025 huggingface dataset:
MicroRNA‐122 protects mice and human hepatocytes from acetaminophen toxicity by regulating CYP1A2 and CYP2E1 expression
Authors: Chowdhary Vivek , Luna Joseph , Teng Kun‐yu , James Laura , Lee William M , Ghoshal Kalpana
Published: November 2023

Although acetaminophen (APAP) is a widely used anti‐pyretic and analgesic, and generally a safe drug, it is a dose‐related toxin in all mammals and causes ~1000 cases of acute liver failure (ALF) annually in the United States. Acetaminophen (APAP) toxicity is a major cause for acute liver failure (ALF) and N‐acetylcysteine (NAC) remains only useful antidote for APAP toxicity till now. MicroRNA‐122 (miR‐122) is a liver‐specific conserved microRNA that regulates metabolic homeostasis and Hepatitis B and C virus replication. It is abundantly expressed in adult mammals and regulates expression levels of many targets in hepatic cells. Although circulating miR‐122 is a sensitive biomarker of ALF, role of this most abundant liver specific miRNA in ALF has not been elucidated. Here, we show that miR‐122 is down regulated in liver biopsies of ALF patients compared to non‐ALF biopsies and in the livers of mice treated with APAP. A dramatic decrease in primary‐miR‐122 expression occurs upon APAP overdose in mice with suppression of its two key trans‐activators, HNF4a and HNF6. More importantly, mortality rate of liver‐specific miR‐122 knockout (LKO) (Mir122loxP/loxP; Alb‐Cre) mice was higher (P<0.0001) than that of the control (Mir122loxP/loxP) mice when injected intraperitoneally with APAP (500mg/kg), which is not lethal to the control mice. Higher basal level of CYP2E1 and CYP1A2 that convert APAP to highly reactive N‐acetyl‐p‐benzoquinone imine (NAPQI) are contributing factors to the sensitivity of LKO mice to APAP overdose. CYP2E1 was upregulated at the protein level whereas Cyp1a2 gene was activated in LKO livers due to upregulation of its key trans‐activator, AHR and the co‐activator MED1. Interestingly, binding of miR‐122 with an intronic site of Ahr primary transcript (hnRNA) and with a 3′‐UTR site of Med1 was identified in the control livers by Argonaute‐HITS‐CLIP analysis, which was validated by the luciferase reporter assay. The loss of these interactions resulted in derepression of Ahr hnRNA and Med1 mRNAs in Mir‐122−/− livers. miR‐122 also suppressed human AHR and MED1 expression by interacting with their 3′‐UTRs. Notably, miR‐122 depletion sensitized differentiated human HepaRG cells to APAP toxicity that correlated with upregulation of AHR, MED1 and CYP1A2 expression. Collectively, these results suggest a critical role of miR‐122 in acetaminophen metabolism and implicate its therapeutic potential in ALF.

Support or Funding Information

This study is supported, in part, by grants R01A193244, R01086978 from NIH and Pelotonia Idea grant.
Tributyrin supplementation attenuates alcohol induced dysbiosis and liver injury
Authors: Donde Hridgandh , Ghare Smita , Wilson Christopher , Gail Cresci , Negy Laura , Gobejishvili Leila , Joshi‐Barve Swati , McClain Craig , Barve Shirish
Published: November 2023

Alcohol‐induced enteric dysbiosis plays an essential role in the pathogenesis of alcoholic liver disease (ALD). Detailed characterization of the alterations in the gut microbiome is needed for understanding their pathogenic role in ALD. Mice were fed liquid Lieber‐DeCarli diet without or with alcohol (5% v/v) for 7 weeks. A subset of mice were administered tributyrin (orally gavaged). Tb is a triglyceride that is rapidly absorbed and metabolized to butyrate by pancreatic lipases. Moreover, it has favorable pharmacokinetics compared with butyrate with low toxicity. Effect of tributyrin on alcohol‐induced intestinal dysbiosis and hepatic injury were evaluated. Metagenomic analysis of the gut microbiome was performed by analyzing the fecal DNA by amplification of the V3–V5 regions of the 16S rRNA gene and large‐scale parallel sequencing on the Illumina MiSeq platform. 7 weeks of chronic alcohol feeding caused a decrease in the diversity of the intestinal microbiome which was reversed by tributyrin administration. Notably, the alcohol‐induced pathogenic changes in the microbiome and the liver were prevented by tributyrin administration. Commensurate with the qualitative and quantitative alterations in the microbiome by butyrate, alcohol‐induced increase in plasma endotoxin and elevation of liver enzymes were significantly reversed by tributyrin. Hence, the increase in the diversity of the bacterial communities of the gut may serve as significant therapeutic target for the prevention/treatment of chronic alcohol intake induced intestinal barrier dysfunction and liver disease.
Monocytic Sphingosine kinase 2 resolves LPS induced lung vascular injury
Authors: Joshi Jagdish C , Rochford Ian C , Baweja Sukriti , Tauseef Mohammad , Chava Koteshwars Rao , Natarajan Viswanathan , Mehta Dolly
Published: November 2023

Acute lung injury (ALI), a leading cause of morbidity and mortality, is characterized by loss of endothelial barrier and neutrophilic injury. While it is known macrophages refold inflammatory signaling but the mechanism by which macrophages resolve microvascular barrier dysfunction during ALI has yet to be identified. We used diphtheria toxin receptor (CD11b‐DTR) promoter driven mice to selectively deplete CD11b+ myeloid population following injury to assess their role in regulating lung vascular injury. Depletion of CD11b+ cells in CD11b‐DTR mice following injury did not exacerbate lung injury but markedly impaired resolution of injury leading to persistent increase in inflammatory cytokine generation. Flow cytometric analysis of CD11b depleted mouse lungs showed a decrease in Ly6c(hi)/SS(low) population while Ly6c(lo)/SS(high) population was increased, suggesting recruited Ly6c(high)/SS(low) monocyte resolves lung vascular injury. Sphingosine‐1‐phosphate (S1P) is a thoroughly studied lipid mediator which facilitates the tightening of the vascular barrier. S1P is generated by the action of sphingosin‐phosphate‐kinase1 (SPHK1) and sphingosine‐phosphate‐kinase2 (SPHK2). We therefore addressed if Ly6c(hi) + cells generates S1P to resolve barrier function and dampens inflammation. We found that adoptive transfer of wild type (WT) or SPHK1−/− CD11b+ bone marrow cells in CD11b depleted mice resolved barrier function while CD11b cells lacking SPHK2 could not. We also determined pro‐inflammatory and anti‐inflammatory cytokine gene expression in bone marrow‐derived macrophages isolated from SPHK1−/−, SPHK2−/− and WT mice after challenging them to LPS for different time point. The IL‐6 expression level was higher in SPHK2−/− BMDM in comparison to WT and SPHK1−/− BMDM. Collectively data suggests that Ly6c(high)/SS(low) in cells is required for restoring vascular barrier function and dampening inflammatory cytokine generation.
Paradoxical Upregulation of Macrophage CXCR7 by Myeloid‐specific Gene Editing in Mice
Authors: Wang Chuan , Alqurashi Thamer , Shen Jianzhong
Published: November 2023

Objective

We previously reported that the chemokine receptor CXCR7 is induced during monocyte‐to‐macrophage differentiation in vitro, leading to a switch of SDF‐1 signaling profile and increased macrophage migration/phagocytosis. However, the role of monocyte/macrophage CXCR7 in vivo remains unknown.

Methods and Results

The CXCR7‐floxed mice (CXCR7‐floxed+/+) were crossed with the LyzM‐Cre mice (LyzM‐Cre+/+) to obtain myeloid‐specific deletion of CXCR7 in vivo. Unlike the systemic knockouts, this new line was viable, fertile, and did not show obvious gross physical or behavioral abnormalities. However, we found significantly higher number of periphery blood monocytes in CXCR7‐floxed+/+/LyzM‐Cre+/+ mice compared with CXCR7‐floxed+/+/LyzM‐Cre−/− mice, indicating a role of CXCR7 in monocytosis. Further study indicated that this phenotype cannot be solely attributed to the disruption of LyzM gene by Cre insertion or the inclusion of floxed sequence to CXCR7 gene. Surprisingly, CXCR7 mRNA and protein expression were dramatically upregulated in CXCR7‐floxed+/+/LyzM‐Cre+/+ mice compared with littermate CXCR7‐floxed+/+/LyzM‐Cre−/− mice, both in bone marrow‐derived macrophages and in peritoneal macrophages, suggesting a potential compensatory mechanism. This is supported by our finding that PCR mapping indicated partial deletion of myeloid CXCR7 gene via the Cre‐LoxP approach and that adenovirus delivery of the Cre gene into CXCR7‐floxed macrophages in vitro mimicked CXCR7 induction in vivo. In addition, in the mouse peritonitis model, injection of thioglycolate led to more macrophage infiltration into the peritoneum in CXCR7‐floxed+/+/LyzM‐Cre+/+ mice.

Conclusions

Myeloid‐specific manipulation of CXCR7 gene by using Cre‐LoxP method leads to compensatory upregulation of CXCR7 in monocytes/macrophages, which is associated with increased monocytosis and cell migration in vivo.

Support or Funding Information

AHA national SDG award
PEA/PLD Association, Reduce Inflammatory Process Associated to Experimental Mouse Model of Atherosclerosis, Induced by Carotid Artery Ligation
Authors: Gugliandolo Enrico , Fusco Roberta , Biundo Flavia , D'Amico Ramona , Di Paola Rosanna , Cuzzocrea Salvatore
Published: November 2023

Atherosclerosis, a disease of the large arteries, is the primary cause of heart disease and stroke. In westernized societies, it is the underlying cause of about 50% of all deaths. Increasing evidence has highlighted the roles of oxidative stress and inflammation in the promotion of atherosclerotic disease. Palmitoylethanolamide (PEA), an endogenous fatty acid amide belonging to the N‐acyl ethanolamine family, has anti‐inflammatory and neuroprotective effects. However, PEA lacks direct capacity to prevent formation of free radicals. Polydatin (PLD) that is a natural precursor of resveratrol has antioxidant activity. Thus, the combination of P.EA and PLD could have beneficial effects on inflammatory process and oxidative stress. The aim of this study was to investigate the effect of PEA in association with PLD in murine model of atherosclerosis induced by carotid artery ligation. This model shows that 14 days after carotid artery ligation there is a significant structural change within the vessel, and that there is an important involvement of the inflammatory pathway in the progression of this disease. In this study we demonstrated that PEA/PLD association treatment reduces atherosclesosis lesion, adhesion molecules (ICAM‐1 (vehicle 7.5 ± 0.7 treatment 2 ± 0.1, p< 0.05), V‐CAM (vehicle 6.5 ± 0.7 treatment 2.5 ± 0.7, p< 0.05)) expression, proinflammatory cytokines (TNF‐α (vehicle 6 ± 0.1 treatment 2 ± 1.41, p< 0.05), IL‐1β (vehicle 7 ± 0.1 treatment 1.5 ± 0.7, p< 0.05)) production, iNOS (vehicle 23914,89 ± 1063,071 treatment 14762,48 ± 3955,329, p< 0.05) and PAR formation (vehicle 7 ± 1.41 treatment 2 ± 0.1, p< 0.05), NF‐kB expression (vehicle 23631,95 ± 7257,002 treatment 275,028 ± 106,1056, p< 0.05) and apoptosis (BAX (vehicle 6 ± 0.1 treatment 1.5 ± 0.7, p< 0.05), Fas‐L (vehicle 22853,57± 959,0419 treatment 12836,61 ± 915,4346, p<0.05)) activation. Our results show that treatment with PEA/PLD 30 mg/Kg is able to reduce vascular damage and attenuates the inflammatory process.
Ubiquitin Proteasome System Inhibition Reduces LRP1 Protein Levels in HepG2 Cells
Authors: Rondón‐Ortiz Alejandro Nico , Gonzales‐Urday Ana Lucia , Martínez‐Málaga Jimena A. , Carr Olivia C. , Maher Timothy J. , Pino‐Figueroa Alejandro
Published: November 2023

Ubiquitin proteasome system (UPS) is a conserved proteolytic process involved in the degradation of damaged and misfolded proteins. Proteins undergoing this event must be conjugated to one or more monomers of ubiquitin, a process known as ubiquitination. UPS inhibition produces protein aggregation which causes cytotoxicity. Low‐density lipoprotein receptor‐related protein‐1 (LRP1) is a transmembrane receptor composed of two subunits, α (515 kDa) and β (85 kDa) located at extracellular and intracellular levels, respectively. This receptor is involved in the uptake of molecules via clathrin for either transcellular transport or lysosomal degradation. UPS inhibition by MG132 increases LRP1 half‐life in HepG2 cells as reported in the literature. Conversely, protein aggregation might have a role in LRP1 reduction.

The purpose of this research was to demonstrate that MG132, a UPS inhibitor, induces cytotoxicity, protein aggregation and reduces LRP1 protein levels in HepG2 cells. These cells were treated for 24 hours with different concentrations of MG132 (0.03 – 3.0 μM) reducing cell viability in a concentration‐dependent manner determined by 3‐(4,5‐dimethylthiazol‐2‐yl)‐5‐(3‐carboxymethoxyphenyl)‐2‐(4‐sulfophenyl)‐2H‐tetrazolium (MTS assay). Whole cell lysates were extracted and separated on SDS‐PAGE. Immunoblotting was used to determine K48‐linked ubiquitinated proteins and LRP1. MG132 at 0.3 and 1 μM increased K48‐linked ubiquitinated proteins and reduced LRP1 protein levels in a concentration‐dependent manner.

These results suggest that UPS inhibition promotes protein aggregation, which can be the cause of cytotoxicity. In addition, MG132 reduced LRP1 protein levels which might be a consequence of protein aggregate accumulation. Further experiments are required to elucidate the LRP1 reduction mechanism when UPS is inhibited.
Prenatal exposure to cyclosporine impair cardiac function in neonates
Authors: El‐Mas Mahmoud , El‐Bassossy Hany , Awan Zuheir
Published: November 2023

Although cardiotoxicity has been recognized as an adverse effect of cyclosporine A (CSA), limited or no information exists regarding effect of prenatal exposure on the cardiac function in the newborn. We tested the hypothesis that prenatal exposure to CSA impair cardiac function in neonates.

Prenatal exposure to CSA (15 mg/kg/day sc) from the day 6 of pregnancy till delivery resulted in a sex‐dependent impairment in ventricular systolic functions in neonates as reflected by a tenfold decrease in ESPVR slope in male neonates while two‐fold decreases in female neonates. In addition, prenatal exposure to CSA led to a significant rise in systolic and diastolic blood pressure in male neonates but not in female neonates. prenatal exposure to CSA caused significant increases in JT interval and tendency to increase in QTc interval in male neonates, but not in females, indicating LV ischemia and arrhythmogenesis. On the other hand, prenatal exposure to CSA resulted in a delay in AV conduction in female neonates, but not in male, rats as reflected by the significant prolongation in P duration and tendency to increase in PR interval

In conclusion, In summary, the current study establishes the importance of the rat sex in defining the magnitude of cardiotoxicity due to CSA prenatal exposure

Support or Funding Information

This work was funded by the Deanship of Scientific Research (DSR), King Abdulaziz University, Jeddah, under grant No. (166 ‐ 110 ‐ D1437).
Probable renin/prorrenin receptor fetal programming in the offspring from rats with pregnancy‐induced hypertension
Authors: Cabrera‐Becerra Sandra Edith , Anguiano‐Robledo Liliana , Gallardo‐Ortiz Itzell A. , Lopez‐Sanchez Pedro
Published: November 2023

It has been described that fetal development is related to health state in adulthood. Barker et. al. have hypothesized that the prevalence of some diseases in adults (as hypertension) are related to intrauterine development, calling this phenomenon as “fetal programming”. Renin angiotensin aldosterone system (RAAS) participates in blood pressure regulation. This system has become more complex due to the finding of new components, such as the prorrenin/renin receptor (PRR), and it is described that some of them might be submitted to fetal programming. The aim of this work was to determine PRR expression in preterm (3rd week of pregnancy), and 1, 4 and 12 weeks old offspring from normal pregnant and pregnancy‐induced hypertensive (PIH) rats. We measured blood pressure, length and body weight and determined PRR and PLZF expression by immunoblot on preterm (3rd. week gestation),1, 4 and 12 weeks old groups. We found lower body weight and length in pups at the third week of gestation, and in 1 and 4 weeks old offspring from PIH. Blood pressure was higher in offspring from PIH rats at 12 weeks. We also found PRR and PLZF expression increased in those groups. These results suggest that PRR might be programmed in fetal stage, and could play a role in development of cardiovascular disease

Support or Funding Information

SIP 20150467 and 20160667
Role of lipocalin‐2 in dietary and aldosterone‐induced renal damages
Authors: Sun Kiwi Wai Yan , Yang Kangmin , Luo Cuiting , Félétou Michel , Villeneuve Nicole , Vanhoutte Paul M , Xu Aimin , Wang Yu
Published: November 2023

Study objective

Both high fat diet and aldosterone challenge can cause significant renal damage, either as a primary cause or as co‐morbidities with other diseases (e.g. cardiovascular diseases and diabetes). Angiotensin‐converting‐enzyme inhibitor (ACEi) and mineralocorticoid receptor inhibitor (MRi; e.g. spinronolactone and eplerenone) are commonly used in these patients. However their mortality is still high and such treatments are not suitable for all patients. Thus, a new therapeutic approach is warranted for renal damages. Herein, we hypothesize that lipocalin‐2 plays a detrimental role in both dietary and aldosterone‐induced renal injuries, and thus could be a novel drug target for therapeutic development.

Methods

Wild‐type (WT), lipocalin‐2 knockout (LKO) and lipocalin‐2 tissue specific knockout in adipose tissue (ADN‐Cre) mice were subjected to eight weeks of high fat diet treatment or to nephrectomy followed by four weeks of aldosterone/nephrectomy/1% saline treatment (ANS). MRi (either spinronolactone or eplerenone) was administered in ANS‐treated groups to compare its efficacy versus lipocalin‐2 deficiency mice in terms of anti‐hypertensive effect, renal damages and fibrosis, and small resistance artery stiffness and tone. Adipose tissue of all mice was also collected for further analysis.

Results

Compared to MRi‐treated wildtype mice, LKO and ADN‐Cre mice showed a better protection against ANS‐induced renal damages, as evidenced by: (1) attenuation of tissue fibrosis, (2) decreased infiltration of macrophages, (3) reduced levels of molecular injury markers, such as kidney injury marker (KIM)‐1, monocyte chemoattractant protein (MCP)‐1, transforming growth factor (TFG)‐β and collagen I. In addition, LKO and ADN‐Cre mice treated with high fat diet showed significant protection from renal damages and reduced adipose tissue fibrosis when compared to controls. As regards small resistance artery stiffness and compliance, ADN‐Cre mice were protected against ANS and high fat diet challenge.

Conclusion

Lipocalin‐2 is involved in the development of the metabolic‐renal syndrome. Targeting its expression and/or signalling pathways could be a novel therapeutic approach to combat these medical complications.

Support or Funding Information

This work was supported by the grants from Hong Kong Research Grant Council [General Research (17121714, HKU 763312 and HKU 780613M), Collaborative Research Fund (C705514G)] and an educational grant from Les Laboratories Servier (RS100503).
An epoxyeicosatrienoic acid analog treated adriamycin‐induced proteinuria and renal fibrosis
Authors: Khan Md Abdul Hye , Goldspink Paul , Imig John D , Falck John R
Published: November 2023

Chronic Kidney Disease (CKD) is a global health concern affecting 16% of the population and an estimated 33 million Americans. Persistent proteinuria and renal fibrosis are the two major contributors in CKD and its progression to end‐stage renal disease. Epoxyeicosatrienoic acid (EET), a CYP epoxygenase metabolite of arachidonic acid, and the EET analogs demonstrated diverse biological actions including renal protection. The aim of this study was to determine if EET‐A, an EET analog, could be used as a treatment for CKD using an adriamycin‐induced kidney injury model. Adriamycin (ADR; 10mg/kg i.v.) was administered to two groups (n=8/group) of male Balb/c mice (8–10 weeks old) whilst the control received saline (i.v.). Urine samples were collected 5 days after ADR injection and 24hr urinary protein excretion was determined. The ADR mice (15.0±4 mg/d) developed elevated proteinuria 5 days after ADR injection compared to control (2.4±0.4 mg/d). After confirming proteinuria 5 days post ADR administration, the ADR mice were treated with vehicle or EET‐A (10mg/kg in drinking water) for two weeks. In ADR mice, EET‐A treatment was able to prevent further proteinuria (11±2 mg/d) compared to the vehicle (25±4 mg/d). EET‐A treatment also reduced renal fibrosis in ADR mice by lowering the collagen positive fibrotic area (1.0±0.1%) compared to vehicle (4.0±1.2%). Overall, the data demonstrated that EET‐A treated ADR‐induced proteinuria and renal fibrosis, and has potential for CKD treatment.

Support or Funding Information

Research Starter Grant to Md. AH Khan from PhRMA Foundation, USA
Discontinuation of Chronic RAAS Inhibitor Therapy at Time of Contrast Media Administration Reduces Degree of Kidney Injury in an Animal Model
Authors: Lauver D. Adam , Parsh Jessica I. , Gurm Hitinder S.
Published: November 2023

It is not known if and when angiotensin‐converting enzyme inhibitors (ACEi) and angiotensin II receptor blockers (ARB) should be discontinued prior to iodinated contrast media (CM) exposure during percutaneous intervention (PCI). Sixteen male New Zealand white rabbits (2.2 – 3.1 kg) were dosed with oral lisinopril or losartan until serum creatinine (SCr) reached steady state. The animals were then administered CM by slow intravenous infusion over 30 min (5.0 g I/kg ioxilan). In half the animals, ACEi or ARB was discontinued on day of CM whereas the other animals were continued on the agents. SCr was measured at time of CM, 1, 2, 24 and 48 hours after CM. Animals were euthanized at 48 hours and kidneys were prepared for histological evaluation. Results were compared to those from 7 control animals who only received CM. SCr stabilized at 7 days after ACEi or ARB initiation. Most (56%) developed contrast‐induced acute kidney injury as defined by SCr increase of greater than 0.5 mg/dl. At 48 hours, SCr values were greater in the animals maintained on ACEi or ARB with a SCr value of 4.03 ± 0.6 (mean ± SEM) in the continued group and 2.24 ± 0.70 in the held group (p‐value = 0.06). Our study demonstrates that continuation of either ACEi or ARB in setting of CM may worsen kidney injury. Further investigation in human subjects of the optimal timing of discontinuation of these agents, in terms of renal protection, prior to CM during PCI or other procedures is needed.
CYP4A/CYP2C Modulation of the Interaction of Calcium Channel Blockers with Cyclosporine on EDHF‐Mediated Renal Vasodilations in Rats
Authors: Hammoud Safaa H , Omar Amal G , Eid Assaad A , El‐Mas Mahmoud M
Published: November 2023

The endothelium‐derived hyperpolarizing factor (EDHF) is a back‐up mechanism that compensates for reduced nitric oxide (NO)/prodtanoids bioavailability. Here we investigated whether (i) cyclosporine (CSA, immunosuppressant drug) upregulates EDHF‐dependent renal vasodilations through altering CYP4A/CYP2C signaling, and (ii) calcium channel blockers modulate the CSA/EDHF/CYP interaction. Rats were treated with CSA, verapamil, nifedipine, or their combination for 7 days. Systolic blood pressure (SBP) was measured by Tail‐cuff plethysmography. Kidneys were then isolated, perfused with physiological solution containing L‐NAME (NOS inhibitor) and diclofenac (cyclooxygenase inhibitor), and preconstricted with phenylephrine. Carbachol vasodilations were established before and after the inhibition of 20‐HETE or EETs. CSA (25 mg.kg−1.day−1 for 7 days) increased SBP and augmented carbachol renal vasodilations. The concurrent administration of verapmil (2 mg.kg−1.day−1) or nifedipine (3 mg.kg−1.day−1) abolished the hypertensive effect of CSA and produced divergent effects on carbachol vasodilations (increases versus decreases). The infusion of MSPPOH (EETs inhibitor) reduced carbachol vasodilations in perfused kidneys of all rat groups, suggesting the importance of EETs in these responses. Moreover, HETE‐20 inhibition by HET0016 increased carbachol vasodilations in control rats, an effect that disappeared in CSA‐treated, and reappeared in rats treated with CSA/verapamil or CSA/nifedipine. The renal protein expression of CYP2C and CYP4A as well as their vasoactive products (EETs and 20‐HETE) were increased in CSA‐treated rats. Whereas the CYP2C/EETs effects of CSA were abolished by verapamil and intensified by nifedipine, the CYP4A/20‐HETE effects were reduced by verapamil only. Overall, CYP2C/EETs and CYP4a/20‐HETE pathways variably modulate the contracting effects of nifedipine and verpamil on CSA‐evoked facilitation of EDHF‐dependent renal vasodilations. These effects may not be related to changes in SBP.

Support or Funding Information

Supported by the Lebanese National Council for Scientific Research (CNRS) Grant Award for Lebanese graduates (Awardee: Safaa Hammoud).
Oroxylin A induces relaxation of porcine basilar arteries via cGMP pathway
Authors: Tsai Andy Po‐Yi , Chen Mei‐Fang , Wang Mei‐Jen , Lee Tony Jer‐Fu
Published: November 2023

The possible involvement of cyclic guanosine monophosphate (cGMP) and potassium channels in mediating oroxylin A (Oro‐A)‐induced relaxation of porcine isolated basilar arteries was examined. Oro‐A, a flavonoid isolated from the natural herb, Scutellariae baicalensis, is used widely for various diseases. Its role in regulating cerebral vascular functions, however, remains unclear. Results from blood vessel myography studies indicated that Oro‐A in a concentration‐dependent manner relaxed U46619 (a thromboxane A2‐receptor agonist)‐contracted basilar arterial rings. The relaxations, which were independent of the endothelial cells and perivascular nerve fibers, were inhibited by blockers for nonspecific potassium channels, preferential calcium‐activated potassium (BKca)‐channels, and preferential ATP‐sensitive potassium (KATP)‐channel. Also, Oro‐A‐induced vasorelaxation was drastically decreased in arteries pre‐contracted by high potassium. By using two‐electrode voltage clamp, Oro‐A, however, did not directly open the BKca‐ or KATP‐channels on oocytes from Xenopus leavis. Moreover, Oro‐A‐induced vasorelaxation was blocked by a soluble guanylate cyclase (sGC) inhibitor, but was not affected by inhibitors of adenylate cyclase or protein kinase A. In addition, concentrations of cGMP but not cAMP in endothelium‐denuded basilar arteries were significantly increased following incubation with Oro‐A. These results suggest that OroA increases synthesis of cGMP which then opens the BKca‐ and KATP‐channels on the smooth muscle cells of the basilar arteries causing relaxation. Together with its reported anti‐inflammatory effects, Oro‐A may be a promising compound for ameliorating cerebral neurovascular dysfunctions.

Support or Funding Information

Supported by Ministry of Science and Technology of Taiwan, Tzu Chi Foundation, and Tzu Chi University
Perivascular Adipose Tissue Modulates the Anticontractile Effect of Cooling in the Rat Aorta
Authors: Rafique Yasmeen , Al‐Bader Maie , Oriowo Mabayoje
Published: November 2023

The perivascular adipose tissue (PVAT) provides mechanical support for blood vessels and exerts an anti‐contractile effect. The present study was designed to investigate how PVAT and gender affect the anti‐contractile effect of cooling in the rat aorta. Aorta segments, with or without PVAT from male and female SD rats were used in this investigation. PE (10−9M – 10−5M) induced concentration‐dependent contractions of aorta segments at 37°C and 24°C. The maximum response and pD2 values were reduced by PVAT in both sexes at 37°C. The reduction in pD2 value was significantly greater in aorta segments from female rats. Cooling (24 °C) reduced the maximum response but not pD2 values to PE in aorta segments without PVAT in both sexes. The anti‐contractile effect of cooling was attenuated in the presence of PVAT. Apocynin (3×10−4M) equipotently inhibited PE‐induced contractions both temperatures. L‐NAME (10−4M) potentiated PE‐induced contractions more in aorta segments without PVAT at both temperatures. The expression of eNOS protein and basal level of nitric oxide (NO) were greater in aorta segments with PVAT at both temperatures. However, PE significantly increased levels of NO only in aorta segments without PVAT. We concluded that PVAT attenuated the anti‐contractile effect of cooling by reducing generation of NO.

Support or Funding Information

Supported by the College of Graduate Studies and a grant (MR 05/15) from Kuwait University Research Sector.
Vascular Endothelial Over‐Expression of Human Soluble Epoxide Hydrolase (Tie2‐sEH Tr) Attenuates Coronary Reactive Hyperemia in Mice: Role of Oxylipins and ω‐Hydroxylases
Authors: Hanif Ahmad , Edin Matthew L , Zeldin Darryl C , Morisseau Christophe , Falck John R , Nayeem Mohammed A
Published: November 2023

Cytochromes P450 metabolize arachidonic acid (AA) into two vasoactive oxylipins with opposing biologic effects: epoxyeicosatrienoic acids (EETs) and omega‐(ω)‐terminal hydroxyeicosatetraenoic acids (HETEs). EETs have numerous beneficial physiological effects, including vasodilation and protection against ischemia/reperfusion injury, whereas ω‐terminal HETEs induce vasoconstriction and vascular dysfunction. We evaluated the effect of these oxylipins on post‐ischemic vasodilation known as coronary reactive hyperemia (CRH). CRH prevents the potential harm associated with transient ischemia. The beneficial effects of EETs are reduced after their hydrolysis to dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolase (sEH). ω‐terminal HETEs are formed by ω‐hydroxylase family members. The relationship among endothelial over‐expression of sEH (Tie2‐sEH Tr), the changes in oxylipins it may produce, the pharmacologic inhibition of ω‐hydroxylases, activation of PPARγ, and CRH response to a brief ischemia is not known. We hypothesized that CRH is attenuated in isolated mouse hearts with endothelial sEH over‐expression through modulation of oxylipin profiles, whereas both inhibition of ω‐hydroxylases and activation of PPARγ enhance CRH. Compared to WT mice, Tie2‐sEH Tr mice had decreased CRH, including repayment volume, repayment duration, and repayment/debt ratio (P < 0.05), whereas inhibition of ω‐hydroxylases increased these same CRH parameters in Tie2‐sEH Tr mice. Inhibition of sEH with t‐AUCB reversed the decreased CRH in Tie2‐sEH Tr mice. Endothelial over‐expression of sEH significantly changed oxylipin profiles, including decreases in DHETs, mid‐chain HETEs, and prostaglandins (P < 0.05). Treatment with rosiglitazone, PPARγ‐agonist, enhanced CRH (P < 0.05) in both Tie2‐sEH Tr and wild type (WT) mice. These data demonstrate that endothelial over‐expression of sEH (through changing the oxylipin profiles) attenuates CRH, whereas inhibition of ω‐hydroxylases and activation of PPARγ enhance it.

Support or Funding Information

This work was supported by National Institutes of Health (HL‐114559) to M. A. Nayeem, and the Intramural Research Program of the NIH, National Institute of Environmental Health Sciences (Z01 ES025034 to D. C. Zeldin).
Microscopic Topography of Atrioventricular Conduction System–A Feature of Potential Clinical Significance
Authors: Sahni Daisy , Randhawa Arpan , Gupta Tulika , Aggarwal Anjali
Published: November 2023

Objectives

Proximity of atrioventricular (AV) conduction system to tricuspid, mitral and aortic valves renders it susceptible to injury leading to disturbances in patients undergoing valve operations and radiofrequency catheter ablations. Hence, the knowledge of topography of AV conduction system and its relations with surrounding cardiac structures is indispensible.

Methods

In 20 cases, a tissue block of 3×4 cm at AV septal junction was excised which included the borders of Koch's triangle. The block was divided into three equal segments by parallel vertical incisions made perpendicular to insertion of septal leaflet of tricuspid valve. Each segment was processed for Hematoxylin & Eosin and Gomori stain.

Results

The upper pole of compact AV node was at the level of mitral valve leaflet in 50% cases. Lower pole of AV node was vertically above tricuspid septal leaflet and at the level of muscular IV septum in 100% and 66.7% cases respectively. The subendocardial depth of upper and lower pole of AV node was 1.45±0.6 mm and 0.92±0.3 mm respectively. Minimum subendocardial depth (0.68±0.2mm) was seen at the center of compact AV node. Penetrating bundle of His was seen at the level of tricuspid septal leaflet just deep to it, while Branching bundle of His was 1.98±1.5 mm below the tricuspid septal leaflet. AV nodal extensions began at the level of mitral valve leaflet while its inferior extension ended 2.92±1.3 mm above tricuspid septal leaflet.

Conclusion

Prior knowledge of distances of AV conduction system from the surrounding landmarks might help in surgical incision planning, safe suture placement and successful device insertion.

Support or Funding Information

From Director PGIMER, Chandigarh, India.
Three Dimensional (3D) Virtual Human Cadaver, Anatomage: An effective teaching tool for Human Anatomy
Authors: Narnaware Raj
Published: November 2023

In an attempt to explore the innovative ways to introduce the knowledge of the human body in terms of its structures and functions, we have first time introduced three dimensional (3D) virtual human cadaver, Anatomage in our human anatomy classes, and discuss whether the use of Anatomage enhances the understanding/knowledge of the human body as determined by their class average with or without using anatomage. We have observed that the use of Anatomage in their teaching significantly increase their class average. Moreover, we also discuss the limitations and advantages of Anatomage as an effective teaching tool to teach human anatomy.
Modification of CUL3 by Related to Ubiquitin (RUB) is partially regulated by the Phytochrome B pathway in Arabidopsis thaliana.
Authors: Orellana Aaron , Christians Matthew
Published: November 2023

Plant growth and development depends on detecting the different wavelengths in the electromagnetic spectrum in order to perceive light conditions and grow accordingly. Phytochrome B, a red and far‐red light photoreceptor in plants, plays an integral role in shade avoidance, flowering time, seed germination, and de‐etiolation. Proper degradation of this photoreceptor and the immediate downstream signaling components, the Phytochrome Interacting Factors (PIFs), is crucial to the function of these processes and occurs via the ubiquitin‐proteasome system. The Light Regulating Bric‐a‐Brac/Tramtrack/Broad Complex proteins (LRB1 and LRB2) interact with CUL3 to form an E3 ligase complex which facilitates the attachment of ubiquitin to phytochrome B and PIF proteins. In plants, cullin‐based E3 ligase activity is dependent on the attachment of RUB (the plant ortholog of NEDD8 in human and other species) to cullin. We found that the LRB proteins contain two highly conserved regions near the N‐terminus which are similar to regions on CUL1/2 and RBX1, and these regions may play a role in the Rubylation process. Further investigation determined that the levels of Rub‐modified CUL3 are modulated by red light, and the PhyB signaling pathway plays a role in this process. To determine if the N‐terminal region of the LRBs play a role in rubylation of CUL3, a truncated version of the LRB gene missing this highly conserved region was inserted into wild type and a lrb1 lrb2 double mutant in Arabidopsis thaliana. Phenotypic, genetic, and immunoblot analysis of homozygous lines featuring the truncated LRBs suggest that the N‐terminal region may affect the ability of the LRBs to regulate red light responses. Future analysis of this region will provide insight into its role in phytochrome degradation and rubylation of CUL3.

Support or Funding Information

This research is funded by an S3 Summer Scholars Grant through the Office of Undergraduate Research at Grand Valley State University
Proprotein Convertase Selectivity in the Activation of the Human Papilloma Virus
Authors: Izaguirre Gonzalo
Published: November 2023

The family of host cell serine proteinases known as proprotein convertases (PCs) is exploited by a variety of human pathogenic viruses to promote viral activation. The PCs, furin, PC5, PACE4, and PC7 reside in the trans‐Golgi network membranes of the constitutive secretion pathway and are expressed in most human tissues. We study PC selectivity for the activation of the human papilloma virus (HPV), and for that purpose we applied an approach that combines the use of recombinant PCs and an assay of HPV cell entry. We found differences in PC selectivity between HPV genotypes 16 and 18 for activation by furin, PACE4, and PC7, and showed that both viral genotypes are inactivated by PC5. These findings imply that proteolytic cleavage of the coat proteins in these two high cancer‐risk HPV genotypes involves multiple cleavage sites with different PC specificity, and challenge the current accepted model of HPV cell entry that postulates that only one PC cleavage site, located on the coat protein L2 at position Arg12, is needed for viral activation. The inactivation by mutagenesis of the six potential PC cleavage site motifs in HPV16, two in L1 and four in L2, demonstrated that additional PC cleavage sites in L1 and L2 are required for HPV16 infection. These results suggest that PC selectivity could be a determinant of viral tropism to anatomical sites of infection based on tissue PC expression profile.
Characterizing small molecule inhibitors of the LINE1 endonuclease
Authors: Smith Michaela , Bertrand Sarah , Bolden Quinteshea , Russell Brittany , DeFreece Cecily
Published: November 2023

Our lab studies the mammalian mobile long interspersed element 1 (LINE1). LINE1 elements mobilize in the genome through a copy and paste itself into new genomic loci through the process of retrotransposition. The full‐length LINE1 element is comprised of two open reading frames, ORF‐1 and ORF‐2. The latter protein, ORF‐2, encodes an endonuclease, which is responsible for nicking DNA near the adenine‐thymine rich base sequence. Proper functioning of LINE1 endonuclease facilitates the mobilization of the element and requires the formation of DNA damage in the form of double‐strand breaks (DSBs), which could, in turn, lead to diseases such as cancer. Approximately eighteen percent of the human genome is comprised of the LINE1 sequence and the expression of LINE1 proteins and mobilization of the LINE1 element has been implicated in several cancers, such as breast and prostate cancers. While events of LINE1 insertion may be mutagenic, the effect of the LINE1 endonuclease on genetic instability is unclear. If the DNA DSBs associated with LINE1 endonuclease are not repaired faithfully, then mutations may result that lead to disease onset or progression. In our laboratory we have identified small molecule inhibitors of the LINE1 endonuclease. Here we present our work investigating the impact of the small molecule inhibitors on the formation of LINE1 endonuclease associated DNA DSBs.

Support or Funding Information

The work presented is supported in part by monies from P20GM103424, TL4GM118968 and 5RL5GM118966‐03 as well as the Louisiana Cancer Research Consortium. We also thank the Molecular and Cellular Biology core at Xavier University of Louisiana.
The Effect of Substrate Presentation and Activation on Neuraminidase NEU2 Specificity
Authors: Woods Robert J , Grant Oliver C , Makeneni Spandana , Foley Bethany L
Published: November 2023

Human exo‐α‐sialidases/neuraminidases cleave terminal sialic acid from glycoconjugates and regulate fundamental cellular processes that are affected by sialic acid recognition, and the disrupted expression of sialidases has been causually linked to a variety of disease states. The activity of cytosolic neuraminidase 2 (NEU2) is not only affected by the linkage type between the terminal sialic acid residue (Neu5Ac in this case) and the penultimate residue in the glycoconjugate (α2‐3/‐6/‐8) linkage), but also by the supramolecular context within which the sialoglycan is presented. It has been shown that NEU2 can cleave Neu5Ac from a mono‐dispersed solution of sialoglycan GM1, but has undetectable activity against GM1 when it is present in a vesicle/micelle. The present study provides a structure‐based rationale for the observed effect of supramolecular organization on NEU2 activity. Data from molecular dynamics simulations of NEU2‐bound glycans illustrate that substrate distortion in the Michaelis complex is likely responsible for linkage specificity.
Antioxidant effects of Agastache mexicana extracts: an in vitro approach
Authors: Esquivel‐Gutiérrez Edgar R. , Coria‐Orozco Emilio , Torres‐Martínez Rafael , Hernández‐García Alejandra , Ríos‐Chávez Patricia , Manzo‐Ávalos Salvador , Saavedra‐Molina Alfredo , Salgado‐Garciglia Rafael
Published: November 2023

Agastache mexicana (Kunth) Lint & Epling (toronjil) is a medicinal plant distributed in the Michoacan state in Mexico and used in traditional medicine for gastrointestinal disorders, anxiety, cardiovascular alterations and efficiently as an anti‐inflammatory and analgesic among others. Ethnobotanical, pharmacological and phytochemical studies have been carry out, however there are currently no scientific relevance studies that support their therapeutic properties. Toronjil have mainly volatile oils (camphene, β‐pinene, limonene, 1,8‐cineole, furfural, citronellal, menthol and pulegone) into their chemical composition, all of them knows as antioxidants. Notwithstanding, there are not reports about the effects of toronjil over the oxidative stress, either in vivo and/or in vitro assays. By this reason, the main aim of this work is to evaluate in vitro the antioxidant properties of Agastache mexicana and some of their isolated compounds. Hydroalcoholic, methanolic and hexanic (Soxhlet) extracts were obtained from toronjil aerial parts and different concentrations (0.001, 0.01, 0.1 and 1 mg ml−1) were tested using the DPPH, ABTS and TBARS methods to evaluate the antioxidant capacity. Our results indicated that the methanol extract (since the 0.1 mg ml−1) has the capacity to scavenger or inhibit the free radicals tested with any assay mentioned here (DPPH, ~93%; ABTS, ~99% and TBARS, ~94%); our GC‐MS analysis reveals the presence of limonene, linalool, mentone, α‐terpineol, pulegone and eugenol mainly. Only eugenol is a potent scavenger of free radicals because its remove ~94%, ~98% and ~98% using the DPPH, ABTS and TBARS methods respectively. The rest of the compounds tested have a relatively low capacity to remove free radicals by either method assayed.

In conclusion, methanolic extract from Agastache mexicana, is a candidate to carry out an “in vivo” approach using an animal model that reflects high oxidative stress.

This work was supported by CIC‐UMSNH/2016‐2017 RSG.

Support or Funding Information

This work was supported by CIC‐UMSNH/2016‐2017 to RSG. CIC‐UMSNH 2.16 to ASM.
The Human Anatomy Laboratory Companion: A Teaching Tool Created by Students, for Students
Authors: Forster Victoria Nicole , Jadeski Lorraine
Published: November 2023

Anatomy is a field that lends itself to a wide range of teaching methodologies depending on student numbers, course level, time constraints, and the level of anatomical detail addressed. The University of Guelph continuously provides high quality human anatomy education to undergraduate students. The program offers dissection‐ and prosection‐based courses at the first, third and fourth year levels of undergraduate study. The combined enrolment in all human anatomy courses is approximately 1000 students per academic year. An ongoing challenge is to continue to provide a very high level of human anatomy education to our students with the available resources. The aim of this project is to develop a ‘companion’ or dissection‐based laboratory manual that: 1) specifically targets curricular goals and course learning objectives, 2) promotes self‐sufficient learning in the laboratory, and 3) increases the efficiency in which material is taught in the laboratory. Fundamental to this laboratory companion is the incorporation of high quality digital images of cadaveric‐based dissections, and has the unique characteristic that it is ‘created by students, for students’. Upper year students who have previously taken the two‐semester dissection‐based course, contribute to the development of the companion, as part of course requirements for a subsequent Teaching, Learning and Knowledge Transfer (TLKT) experience. Thus, these students (TLKT) work closely with faculty and graduate students to prepare detailed, precise and step‐wise dissections that are captured digitally, and incorporated into this resource. Significant progress has been made in developing the framework for the companion, as dissections and digital images have been created. Preliminary data has been collected from students during the Winter 2017 semester of the third year course, with the use of questionnaires (n=305). The results show that this educational tool is desired by students to help them prepare for and review laboratory course content and excel independently. Subsequent to the development of the companion, it will be implemented as a learning resource in the dissection (HK*3401) and prosection (HK*3501) courses at the University of Guelph. The pedagogical impact will be investigated through comparison with historic controls and will be measured using student performance and student perception of the course experience as outcomes.