bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Influence of Covid‐19‐Based mRNA Vaccines on Measures of Conduit Artery and Microvascular Endothelial Function
Authors: Serviente Corinna , Matias Alexs , Erol Muhammet E. , Calderone Mia , Layec Gwenael 
Published: May 2022

With the emergence of severe acute respiratory syndrome 2 (SARS‐CoV‐2), leading to coronavirus disease 19 (COVID‐19), a new class of messenger RNA (mRNA) vaccines have been broadly used. These vaccines have been associated with inflammatory sides effects such as myo‐ and peri‐carditis. Despite the well‐known link between inflammation and endothelial dysfunction, it is unknown whether these vaccines may transiently impair endothelial function, and therefore increase cardiovascular risk. Aim: To evaluate the influence of COVID‐19‐based mRNA vaccines on macro‐ and micro‐vascular endothelial function in healthy adults. We hypothesized that macro‐ and micro‐vascular endothelial function would be reduced following vaccination.MethodsPrior to, and ~48 hrs following the second dose of COVID‐19‐based mRNA vaccines, brachial artery flow‐mediated dilation (FMD, macrovascular function), reactive hyperemia (microvascular function), and cutaneous intradermal microdialysis coupled with laser Doppler flowmetry (microvascular function) were measured in healthy young adults. For microdialysis, a standard local heating protocol (42⁰C) was utilized, followed by infusion of the Nitric Oxide Synthase inhibitor Nω‐Nitro‐L‐arginine methyl ester hydrochloride (L‐NAME, 15mM), and induction of maximum vasodilation (43⁰C+28mM Sodium Nitroprusside). Sites included: control (lactated Ringer’s), ascorbic acid (10 mM), l‐arginine (10 mM), and tetrahydrobiopterin (BH4,10 mM). C‐reactive protein (CRP), a marker of systemic inflammation, was measured from fasting serum samples at both time points.ResultsVaccination (mRNA‐1273, n=3 and BNT162b2, n=6, 5men/4women, 34±2yrs) induced an increase in CRP (pre: 0.2±0.1mg/dL vs. post: 2.1±0.5mg/dL, p=0.008, r‐statistic=‐0.89), but had no effect on FMD (pre: 3.6±0.9% vs. post: 4.6±1.1%, p=0.43, Cohen’s d=0.28) or reactive hyperemia (pre: 20178±3236 AU vs. post: 28426±4573 AU, p=0.14, Cohen’s d=0.55). There was no association between the change in FMD (r=‐0.07, p=0.87) or reactive hyperemia (r=‐0.48, p=0.19) and the change in CRP. Vaccination also had no effect on microvascular nitric oxide (NO)‐dependent dilation (pre: 77.0±2.5% vs post: 77.3±2.4%, p=0.91, ηp2=0.002), nor was there an effect of microdialysis treatment on NO‐dependent dilation (p=0.74, ηp2=0.05). Following vaccination, the difference in NO‐dependent dilation relative to control was unaffected by treatment (p=0.25, η2=0.11) with no difference in the effects of BH4 (6.3±4.9%), l‐arginine (8.3±5.2%), or ascorbic acid (‐2.6±4.2%, all p>0.05). The change in NO‐dependent dilation at the control site was not related to the change in CRP (r=0.32, p=0.40).ConclusionsThere was a significant inflammatory response to COVID‐19‐based mRNA vaccines ~48 hrs following the second vaccine dose. Despite this, macro‐ and micro‐vascular endothelial function and NO‐dependent dilation were preserved in healthy adults.
Metformin down‐regulates TGF beta signal transduction and production of PAR2 N‐terminus cleaving protease activity in an NR4a1 dependent manner in a PC3 prostate cancer cell line
Authors: Mihara Koichiro , Saifeddine Mahmoud , Hollenberg Morley D. 
Published: May 2022

OBJECTIVEOur aim is to understand the mechanisms of metformin’s antitumor actionsHYPOTHESISMetformin is known to have antitumour effects in cancers like head & neck cancer, that may involve its regulation of TGF‐β signalling, but the metformin receptor mechanisms for this action are not known. We recently discovered that the action of metformin to preserve hypoglycaemia‐impaired endothelial cell function regulated by proteinase‐activated receptor‐2 (PAR2) requires its interaction with the orphan nuclear receptor, NR4A1 (PubMed ID 34452975). PAR2 also plays a role in regulating prostate cancer PC3 cell function whereby autocrine signaling occurs in PC3 cells which secrete a PAR2‐cleaving/activating matrix metalloproteinase, MMP2. We hypothesized that metformin can act in PC3 prostate cancer‐derived cells via NR4A1 to change TGF‐β signalling and to alter the production of PAR‐cleaving proteinases.METHODSThe ability of TGF‐β to upregulate connective tissue growth factor (CTGF) mRNA in PC3 cells was measured for metformin treated cells (</=100 μM) that were transfected with NR4A1, C‐terminally tagged with red‐fluorescent protein (RFP): either wild‐type (WT) or mutant (PPGG) murine NR4A1 that does not interact with metformin. TGF‐β upregulated CTGF mRNA in PC3 cells was monitored after metformin treatment for 20 hours. In similarly transfected WT and mutant PPGG NR4A1‐expressing PC3 cells, treated or not with metformin (</=100 μM), the cleavage status of dually tagged transfected N‐terminal RFP/C‐terminal GFP‐PAR2 was monitored: (confocal imaging: yellow = intact receptor; green = cleaved receptor). Concurrently, the abundance of MMP activity of PC3 cell supernatants was measured by a fluorogenic MMP substrate, Mca‐RPKPVE‐Nval‐WRK(Dnp)‐NH2 (ES002).RESULTSThe expected TGF‐β upregulated CTGF mRNA in PC3 cells was prevented by metformin treatment, but only for cells transfected with the WT and not the mutant PPGG NR4A1. Metformin acted at concentrations (</=100 μM) far lower than those that affect mitochondrial complex I (> mM). We also found that in WT, but not PPGG mutant‐transfected PC3 cells, metformin (</=100 μM) down‐regulated MMP PAR2‐cleaving proteinase activity.CONCLUSIONSOur results show that low concentrations of metformin (</=100 μM), in the presence of wild‐type but not PPGG mutant NR4A1 which does not enable metformin interactions at the NR4A1 alternative ligand binding site, can both block TGF‐ β signaling and reduce cell‐secreted MMP activity that in turn can drive tissue invasion and metastasis via PAR2 activation. Thus, NR4A1 may play a role in the antitumour actions of metformin.
Nitric Oxide Therapy Attenuates Heart Failure with Preserved Ejection Fraction
Authors: Koul Kashyap , LaPenna Kyle , Li Zhen , Doiron Jake , Wang John , Sharp Thomas E. , Lefer David J. 
Published: May 2022

IntroductionNitric oxide (NO) based therapeutics have proven to exert cardioprotective actions in the setting of heart failure with reduced ejection fraction (HFrEF). However, NO therapy has failed to exert beneficial effects in clinical studies of heart failure with preserved ejection fraction (HFpEF).We investigated (1) the effects of the NO donor, sodium nitrite, as a monotherapy, (2) hydralazine monotherapy, a known vasodilator and antioxidant, and (3) combination therapy with both drugs in a murine model of HFpEF. We assessed left ventricular (LV) function, oxidative/nitrosative stress, exercise tolerance, and cardiac fibrosis.Materials and MethodsMale, C57/BL6N (n=15 per group) were placed on a Western high fat diet (60% kCal from fat) and treated with L‐NG‐Nitro arginine methyl ester (L‐NAME, 0.5 g/L/day) in the drinking water beginning at 10 weeks of age. At 15 weeks of age, mice were randomly assigned to four separate groups for five additional weeks: HFpEF control, sodium nitrite in drinking water (75 mg/L), hydralazine (2 mg/kg/day, i.p., b.i.d.), or the combination of sodium nitrite and hydralazine.Plasma was collected to measure circulating 8‐isoprostane and 3‐nitrotyrosine levels, biomarkers of oxidative and nitrosative stress respectively. In addition, cardiac tissue was harvested at 20 weeks for Masson’s Trichrome staining and to measure cardiac nitrite levels. Echocardiography and exercise tolerance testing was performed at 10, 15, and 20 weeks of age.ResultsThere was a significant increase in circulating nitrite levels (p < 0.05) in the sodium nitrite + hydralazine group compared to the control and either monotherapy groups. There was no significant difference in the cardiac tissue nitrite levels between the combination therapy and the control group. Plasma 8‐isoprostane and 3‐nitrotyrosine levels were significantly (p < 0.05) reduced in the sodium nitrite + hydralazine group compared to the HFpEF control group, indicating a reduction in oxidative and nitrosative stress. Although there was no significant difference in the extent of interstitial cardiac fibrosis between groups, there was a significant decrease (p < 0.05) in myocardial perivascular fibrosis in the sodium nitrite + hydralazine group compared to control. Left ventricular end‐diastolic pressure (LVEDP) was significantly reduced in all treatment groups, and LVEDP was reduced to the greatest extent in the nitrite + hydralazine group vs. HFpEF control. Furthermore, we observed a significant (p < 0.01) improvement in LV diastolic function (i.e., E/e’) in the sodium nitrite and hydralazine group that was superior to either of the treatments alone. There was no significant difference in the exercise distance or duration among the various groups.ConclusionThese data demonstrate that the combination of a powerful antioxidant with NO donor therapy significantly enhances the beneficial effects NO therapy in the setting of severe HFpEF.
Differential Regulation of Lhx9 Expression in Mouse and Chicken Limbs
Authors: Kim Jean Young , Britton Jessica C. , Pira Charmaine U. , Oberg Kerby C. 
Published: May 2022

The distal tip of the developing vertebrate limb is covered by a rim of thickened ectoderm called the apical ectodermal ridge (AER). The AER secretes fibroblast growth factors (Fgfs) into the underlying mesoderm to control outgrowth and proximal‐distal patterning. Patterning during outgrowth proceeds from proximal to distal, with the proximal segment or stylopod being laid down first, then the zeugopod, and the distal segment or autopod forming last. Secreted Fgfs from the AER maintain a sub‐AER population of undifferentiated mesoderm, called the progress zone, that provides cells for this progressive differentiation.In mice, Lhx9and its homologue Lhx2are expressed in the sub‐AER mesoderm and appear to play redundant roles in maintaining the expression of other patterning molecules, such as Shh. Interestingly in the chicken, the expression of LHX9 is restricted to the anterior aspect of the sub‐AER mesoderm, while LHX2 retains the full distal expression seen in mice. Recent clinical reports of an LHX9 missense mutation in a patient with hypoplastic/missing thumbs and great toes suggest that LHX9 can play a critical role in normal limb development. We hypothesized that the chicken cis‐regulatory modules (CRMs) regulating Lhx9 might have subtle species‐specific sequence variations compared to the murine CRMs that correlate with this differential expression pattern.To test this hypothesis, potential CRMs were identified near the Lhx9 locus by in silicoanalysis, using sequence conservation and regulatory chromatin markers from published ChIP‐seq data. The potential CRMs were isolated and inserted into CRM‐reporter constructs. The limb‐related activity of a potential CRM was then determined by targeted regional electroporation into the distal chicken limb mesoderm.We identified a highly conserved, transcription‐factor‐rich 623 bp sequence within intron 12 of DENDD1B 132 kbp upstream of the chicken LHX9 transcription start site. The potential CRM was associated with H3k4me2, H3k27ac, p300 and H3k27me3. The associated CRM‐reporter construct displayed activity in both anterior and posterior sub‐AER limb mesoderm overlapping LHX9 and LHX2 expression. These findings are consistent with a distal sub‐AER LHX9enhancer, but do not account for the lack of posterior expression. This suggests that additional CRMs contribute to the final expression pattern of LHX9 in the limb. Twenty‐two additional potential CRMs were identified and will be further characterized to determine their role in restricting LHX9expression from the posterior mesoderm in chickens. Characterizing the differential regulation of Lhx9 during limb development may provide insight into the species‐specific variation of limb patterning.
Fascicular Organisation and Neuroanatomy of the Porcine and Human Vagus Nerves: Allowing for Spatially Selective Vagus Nerve Stimulation
Authors: Thompson Nicole , Ravagli Enrico , Mastitskaya Svetlana , Iacoviello Francesco , Perkins Justin , Shearing Paul , Aristovich Kirill , Holder David 
Published: May 2022

Vagus nerve stimulation (VNS) often results in off‐target effects due to the unknown fascicular organisation of the cervical vagus nerve and thus nonspecific nature of stimulation. This currently limits its therapeutic efficacy in treating epilepsy and depression and restricts its extension to treating numerous other conditions, such as heart failure and lung injury, for which it could be a promising treatment. The purpose of this work was to determine the fascicular anatomical representation of the recurrent laryngeal, cardiac and pulmonary branches of the vagus nerve at the mid‐cervical level of VNS cuff placement in the vagus nerve in pigs using micro‐computed tomography (microCT) to trace anatomical connections to end‐organs with the intention of extension to humans.The left vagus nerves were dissected from mid‐cervical level to the branches of the lungs, heart and larynx in four domestic female 60‐70 kg pigs. Nerves were fixed, sectioned into 4 cm lengths with registration landmarks, soaked in Lugol’s solution (1% total iodine) for five days for sufficient contrast staining and microCT‐scanned (Nikon XT H 225, Nikon Metrology) with a molybdenum target and 7 µm voxel size, 35 kVp energy, 114 mA current, 4 W power, 0.25 fps in 4 s exposure time, and 3176 projections. Subsequently, scans were reconstructed in CT Pro 3D (Nikon Metrology) and analysed in ImageJ. Fascicles of the three target organs/fascicle groups were segmented and traced with Neurolucida 360 (MBF Bioscience) from the point the organ‐specific branch enters the vagus nerve to mid‐cervical level (20‐25 cm total length per nerve).The three identified fascicular groups remained discrete at the cervical level (26%, 23% and 4% of total vagus nerve area for pulmonary, recurrent laryngeal and cardiac fascicle groups, respectively, with significant separation of centres of mass (P<0.05)). These three distinct regions contained overlap only between a small number of pulmonary and recurrent laryngeal fascicles (<11%). The cardiac fascicles remained separate. This confirms organotopic organisation of the cervical vagus nerve according to organ/function for cardiac, pulmonary and recurrent laryngeal fascicles. Work is in progress to delineate similar fascicular organisation in excised cadaveric human vagus nerves. Knowledge of the fascicular organisation of the vagus nerve will enable accurate spatially selective VNS which may allow for avoidance of off‐target effects and so improve its therapeutic efficacy.
The Impact of HIV Infection in Cocaine Seeking Behaviors
Authors: Xie Qiaowei , Jackson Joshua , Barker Jacqueline 
Published: May 2022

BackgroundSubstance use disorders (SUDs) are characterized by high propensity to relapse and are highly comorbid with HIV infection. The underlying neurobiology of this relationship is poorly understood. Preclinical research on the neurobiobehavioral outcomes of progressive HIV infections may yield crucial information to improve SUD prognosis and reduce risk of relapse in people living with HIV.MethodsTo model progressive HIV, adult male and female C57BL6J mice were inoculated with EcoHIV, a chimeric HIV‐1 which infects murine cells by replacing envelope glycoprotein gp120 with gp80. Five weeks after inoculation mice were trained in a cocaine conditioned place preference (CPP) paradigm (10mg/kg, i.p.) followed by extinction training. Stress‐induced reinstatement was assessed following administration of yohimbine, an α2 adrenergic antagonist, to determine EcoHIV effects. Neuronal activation in the prefrontal cortex (PFC) and nucleus accumbens (NAc) by cocaine exposure was assessed by quantitative analysis the expression of the immediate early gene, cFos.ResultsEcoHIV and sham‐inoculated mice exhibited similar cocaine CPP and extinction. However, EcoHIV mice showed significantly increased yohimbine‐induced reinstatement rate (p<0.05). cFos analysis revealed an elevated neural activation in PFC and NAc in response to cocaine. However, only neurons within infralimbic PFC (IL) and NAc shell subregions showed further increases in activity in cocaine exposed mice infected with EcoHIV vs shams.ConclusionEcoHIV infected mice escalated stress‐induced reinstatement of cocaine seeking compared to controls, suggesting that people living with HIV may be at elevated risk for stress‐induced relapse. Progressive EcoHIV infection may dysregulate IL and NAcShell activity to facilitate the cocaine seeking and reinstatement. Ongoing research will characterize how EcoHIV infection dysregulate IL‐NAcShell pathway specific activity during cocaine reward learning and stress‐induced relapse.
Synchronous biventricular pressure‐volume loops in rodents using a closed‐chest approach
Authors: West Christopher R. , Ahmadian Mehdi , Wainman Liisa 
Published: May 2022

IntroductionThe assessment of left‐ and right‐sided cardiac function via closed‐chest pressure‐volume loops is important to understanding and diagnosing a wide range of cardiovascular pathologies. In the vast majority of pre‐clinical studies, either left‐ or right‐sided heart function is assessed in isolation, or, more recently, by sequentially right‐ and left‐sided catheterization1. Synchronous biventricular catheterization would offer a significant advancement for pre‐clinical research. To date, closed‐chest synchronous biventricular pressure‐volume loops have only been obtained in a few studies, limited to porcine models2. Here, we describe initial results from what we believe to be the first synchronous biventricular pressure‐volume loops obtained in the rodent using a closed‐chest approach.AimTo obtain biventricular pressure‐volume loops in the anesthetized rat using a closed‐chest approachMethodsA total of 8 Wistar rats were anesthetized with urethane (1.5mg/kg), tracheotomised, ventilated, and subsequently had a solid state catheter placed in the femoral artery to measure blood pressure, and admittance pressure‐volume catheters placed in the right‐ and left‐ventricle to obtain synchronous biventricular pressure‐volume loops. Admittance probes were set to different frequencies to avoid cross‐talk. All catheters were left to stabilize for a minimum of 10 min after which we extracted and averaged 1 minute of basal pressure‐volume indices.ResultsReliable ventricular pressure waveforms were obtained from both the left (i.e., left‐ventricular end‐systolic pressure = 106±12mmHg) and right (i.e., right‐ventricular end systolic pressure = 28±8mmHg) side in all eight animals, and we were able to obtain standard left‐sided pressure‐volume loops from all eight animals (i.e., stroke work = 22±5mmHg/ml). We were only able to obtain reliable right‐sided pressure‐volume loops in two of eight animals, due to poor volume signals in the remaining 6 animals. In the two animals we were able to obtain synchronous pressure‐volume loops there was clear evidence of ventricular dependence across the respiratory cycle (Fig.1).ConclusionsObtaining synchronous biventricular pressure‐volume loops using a closed‐chest approach in rats is possible but technically challenging. Further work is needed to better obtain right‐sided ventricular volumes.References

Potus F. et al., J. Vis. Exp (160), e61088, doi:10.3791/61088 (2020).
Lyhne MD. et al., J. Vis. Exp.(171), e62661, doi:10.3791/62661 (2021).
The role of Kisspeptin, KISS1 and KISS1R genes in primary female infertility: a case‐control study of a Nigerian population
Authors: Okafor Izuchukwu A. , Saanu Oluseun O. , Olayemi Oladapo , Omigbodun Akinyinka 
Published: May 2022

IntroductionThere is growing evidence that genetic changes are present in as many as 10% of infertile females. The Kisspeptin system plays a critical regulatory role in female reproductive functions. However, the extent of its role is not yet evident in primary infertility.ObjectiveThis study investigated the relative gene expression levels of Kisspeptin, KISS1, and KISS1R in primary infertile and fertile Nigerian females.Materials and MethodsThis is a case‐control study of 50 fertile and 54 primary infertile Nigerian females (20 to 40 years) attending the gynecology and family planning clinics of University College Hospital, Nigeria, and met the study’s inclusion criteria between August and November 2020. Basic clinical and sociodemographic information was collected from each participant using a clinical proforma. Five ml of blood samples were also collected by venepuncture from both the cases and control subjects for Kisspeptin, KISS1, and KISS1R genes analysis, using ELISA and RT‐qPCR techniques.ResultsBMI of cases was significantly different from the study controls, though both are still in the overweight category (27.98±0.87 vs. 28.18±0.59; p=0.004). Menarcheal age was not significantly different (p=0.573) but shows a significant strong association with ovarian factor infertility (p=0.008, χ2=13.839, φc= 0.458) and tubal factor infertility (p=0.025, χ2=11.111; φc=0.527). There is no change in the relative expression of Kisspeptin, Kiss1, and Kiss1R genes (P>0.05). However, Kisspeptin levels in primary infertile women showed a weak negative correlation (r = ‐.305; p=0.039) and a good predictive model for Kiss1 gene expression (p=0.001; r=0.458).ConclusionFemales with primary infertility may present with overweight in no significantly different way than fertile females. Changes in the Kisspeptin levels and relative expressions of Kiss1 and Kiss1R may not be useful in diagnosing primary infertility. Menarcheal age should be investigated as an additional diagnostic indicator for tubal and ovarian factor infertility for primary infertile women.SignificanceChanges in the Kisspeptin system alone may not explain the primary female infertility due to the complexity of the disease. A look into the role of the Kisspeptin system in the specific causative factors of primary female infertility could be more revealing.