Smooth Contraction Research Articles

Transcriptomic Analysis Reveals Intrinsic Abnormalities in Endometrial Polyps.

Endometrial polyps (EPs) are benign overgrowths of the endometrial tissue lining the uterus, often causing abnormal bleeding or infertility. This study analyzed gene expression differences between EPs and adjacent endometrial tissue to elucidate intrinsic abnormalities promoting pathological overgrowth. RNA sequencing of 12 pairs of EPs and the surrounding endometrial tissue from infertile women revealed 322 differentially expressed genes. Protein-protein interaction network analysis revealed significant alterations in specific signaling pathways, notably Wnt signaling and vascular smooth muscle regulation, suggesting these pathways play critical roles in the pathophysiology of EPs. Wnt-related genes DKK1 and DKKL1 were upregulated, while GPC3, GREM1, RSPO3, SFRP5, and WNT10B were downregulated. Relevant genes for vascular smooth muscle contraction were nearly all downregulated in EPs, including ACTA2, ACTG2, KCNMB1, KCNMB2, MYL9, PPP1R12B, and TAGLN. Overall, the results indicate fundamental gene expression changes promote EP formation through unrestrained growth signaling and vascular defects. The intrinsic signaling abnormalities likely contribute to clinical symptoms of abnormal uterine bleeding and infertility common in EP patients. This analysis provides molecular insights into abnormal endometrial overgrowth to guide improved diagnostic and therapeutic approaches for this troublesome women's health condition. Confirmation of expanded cohorts and further investigations into implicated regulatory relationships are warranted.

THE EFFECTIVENESS OF GINGER (Zingiber officinale) AS AN ANTIEMETIC ON PREGNANT WOMEN

Nausea and vomiting during pregnancy are common occurrences in the first trimester. This can be attributed to an increase in estrogen and progesterone hormones produced by human chorionic gonadotropin (HCG) in the placenta. Various therapies can be employed as antiemetics in pregnant women, and one of them is traditional therapy involving the administration of ginger extract. This research design uses a literature review methodology, gathering data from international and national journals that meet the inclusion and exclusion criteria. The databases utilized include Google Scholar, PubMed, and Garuda. To understand the effectiveness of ginger extract administration as an antiemetic in pregnant women. The methodology employed is a literature review, utilizing the Google Scholar database to search for articles published between 2013 and 2023 in both Indonesian and English languages. Ginger can reduce nausea and vomiting by modulating gastrointestinal effects such as smooth muscle contractions and stimulating gastric motility. It inhibits 5-HT3 receptors in the central nervous system, hampers carminative effects, reduces cisplatin effects, and has effects similar to dimenhydrinate. Administering ginger infusion twice a day for 4-7 days has shown a decrease in the occurrence of nausea and vomiting in pregnant women. Additionally, ginger aromatherapy with 5 ml for four consecutive days can reduce the incidence of nausea and vomiting in pregnant women with hyperemesis gravidarum. Ginger extract can reduce nausea and vomiting in pregnant women.

The BK Channel Limits the Pro-Inflammatory Activity of Macrophages.

Macrophages play a crucial role in the innate immune response, serving as key effector cells in the defense against pathogens. Although the role of the large-conductance voltage and calcium-activated potassium channel, also known as the KCa1.1 or BK channel, in regulating neurotransmitter release and smooth muscle contraction is well known, its potential involvement in immune regulation remains unclear. We employed BK-knockout macrophages and noted that the absence of a BK channel promotes the polarization of macrophages towards a pro-inflammatory phenotype known as M1 macrophages. Specifically, the absence of the BK channel resulted in a significant increase in the secretion of the pro-inflammatory cytokine IL-6 and enhanced the activity of extracellular signal-regulated kinases 1 and 2 (Erk1/2 kinases), Ca2+/calmodulin-dependent protein kinase II (CaMKII), and the transcription factor ATF-1 within M1 macrophages. Additionally, the lack of the BK channel promoted the activation of the AIM2 inflammasome without affecting the activation of the NLRC4 and NLRP3 inflammasomes. To further investigate the role of the BK channel in regulating AIM2 inflammasome activation, we utilized BK channel inhibitors, such as paxilline and iberiotoxin, along with the BK channel activator NS-11021. Pharmacological inactivation of the BK channel increased, and its stimulation inhibited IL-1β production following AIM2 inflammasome activation in wild-type macrophages. Moreover, wild-type macrophages displayed increased calcium influx when activated with the AIM2 inflammasome, whereas BK-knockout macrophages did not due to the impaired extracellular calcium influx upon activation. Furthermore, under conditions of a calcium-free medium, IL-1β production following AIM2 inflammasome activation was increased in both wild-type and BK-knockout macrophages. This suggests that the BK channel is required for the influx of extracellular calcium in macrophages, thus limiting AIM2 inflammasome activation. In summary, our study reveals a regulatory role of the BK channel in macrophages under inflammatory conditions.

Direct modulation of TRPC ion channels by Gα proteins.

GPCR-Gi protein pathways are involved in the regulation of vagus muscarinic pathway under physiological conditions and are closely associated with the regulation of internal visceral organs. The muscarinic receptor-operated cationic channel is important in GPCR-Gi protein signal transduction as it decreases heart rate and increases GI rhythm frequency. In the SA node of the heart, acetylcholine binds to the M2 receptor and the released Gβγ activates GIRK (I(K,ACh)) channel, inducing a negative chronotropic action. In gastric smooth muscle, there are two muscarinic acetylcholine receptor (mAChR) subtypes, M2 and M3. M2 receptor activates the muscarinic receptor-operated nonselective cationic current (mIcat, NSCC(ACh)) and induces positive chronotropic effect. Meanwhile, M3 receptor induces hydrolysis of PIP2 and releases DAG and IP3. This IP3 increases intracellular Ca2+ and then leads to contraction of GI smooth muscles. The activation of mIcat is inhibited by anti-Gi/o protein antibodies in GI smooth muscle, indicating the involvement of Gαi/o protein in the activation of mIcat. TRPC4 channel is a molecular candidate for mIcat and can be directly activated by constitutively active Gαi QL proteins. TRPC4 and TRPC5 belong to the same subfamily and both are activated by Gi/o proteins. Initial studies suggested that the binding sites for G protein exist at the rib helix or the CIRB domain of TRPC4/5 channels. However, recent cryo-EM structure showed that IYY58-60 amino acids at ARD of TRPC5 binds with Gi3 protein. Considering the expression of TRPC4/5 in the brain, the direct G protein activation on TRPC4/5 is important in terms of neurophysiology. TRPC4/5 channels are also suggested as a coincidence detector for Gi and Gq pathway as Gq pathway increases intracellular Ca2+ and the increased Ca2+ facilitates the activation of TRPC4/5 channels. More complicated situation would occur when GIRK, KCNQ2/3 (IM) and TRPC4/5 channels are co-activated by stimulation of muscarinic receptors at the acetylcholine-releasing nerve terminals. This review highlights the effects of GPCR-Gi protein pathway, including dopamine, μ-opioid, serotonin, glutamate, GABA, on various oragns, and it emphasizes the importance of considering TRPC4/5 channels as crucial players in the field of neuroscience.

ATP-Induced Contractile Response of Esophageal Smooth Muscle in Mice.

The tunica muscularis of mammalian esophagi is composed of striated muscle and smooth muscle. Contraction of the esophageal striated muscle portion is mainly controlled by cholinergic neurons. On the other hand, smooth muscle contraction and relaxation are controlled not only by cholinergic components but also by non-cholinergic components in the esophagus. Adenosine triphosphate (ATP) is known to regulate smooth muscle contraction and relaxation in the gastrointestinal tract via purinergic receptors. However, the precise mechanism of purinergic regulation in the esophagus is still unclear. Therefore, the aim of the present study was to clarify the effects of ATP on the mechanical responses of the esophageal muscle in mice. An isolated segment of the mouse esophagus was placed in a Magnus's tube and longitudinal mechanical responses were recorded. Exogenous application of ATP induced contractile responses in the esophageal preparations. Tetrodotoxin, a blocker of voltage-dependent sodium channels in neurons and striated muscle, did not affect the ATP-induced contraction. The ATP-evoked contraction was blocked by pretreatment with suramin, a purinergic receptor antagonist. RT-PCR revealed the expression of mRNA of purinergic receptor genes in the mouse esophageal tissue. The findings suggest that purinergic signaling might regulate the motor activity of mouse esophageal smooth muscle.

Transcriptomic Analysis Provides Insights into Candidate Genes and Molecular Pathways Involved in Growth of Mytilus coruscus Larvae.

Growth is a fundamental aspect of aquaculture breeding programs, pivotal for successful cultivation. Understanding the mechanisms that govern growth and development differences across various stages can significantly boost seedling production of economically valuable species, thereby enhancing aquaculture efficiency and advancing the aquaculture industry. Mytilus coruscus, a commercially vital marine bivalve, underscores this importance. To decipher the intricate molecular mechanisms dictating growth and developmental disparities in marine shellfish, we conducted transcriptome sequencing and meticulously analyzed gene expression variations and molecular pathways linked to growth traits in M. coruscus. This study delved into the molecular and gene expression variations across five larval development stages, with a specific focus on scrutinizing the differential expression patterns of growth-associated genes using RNA sequencing and quantitative real-time PCR analysis. A substantial number of genes-36,044 differentially expressed genes (DEGs)-exhibited significant differential expression between consecutive developmental stages. These DEGs were then categorized into multiple pathways (Q value < 0.05), including crucial pathways such as the spliceosome, vascular smooth muscle contraction, DNA replication, and apoptosis, among others. In addition, we identified two pivotal signaling pathways-the Hedgehog (Hh) signaling pathway and the TGF-beta (TGF-β) signaling pathway-associated with the growth and development of M. coruscus larvae. Ten key growth-related genes were pinpointed, each playing crucial roles in molecular function and the regulation of growth traits in M. coruscus. These genes and pathways associated with growth provide deep insights into the molecular basis of physiological adaptation, metabolic processes, and growth variability in marine bivalves.

Stroke Alters the Function of Enteric Neurons to Impair Smooth Muscle Relaxation and Dysregulates Gut Transit.

Gut dysmotility is common after ischemic stroke, but the mechanism underlying this response is unknown. Under homeostasis, gut motility is regulated by the neurons of the enteric nervous system that control contractile/relaxation activity of muscle cells in the gut wall. More recently, studies of gut inflammation revealed interactions of macrophages with enteric neurons are also involved in modulating gut motility. However, whether poststroke gut dysmotility is mediated by direct signaling to the enteric nervous system or indirectly via inflammatory macrophages is unknown. We examined these hypotheses by using a clinically relevant permanent intraluminal midcerebral artery occlusion experimental model of stroke. At 24 hours after stroke, we performed invivo and exvivo gut motility assays, flow cytometry, immunofluorescence, and transcriptomic analysis. Stroke-induced gut dysmotility was associated with recruitment of muscularis macrophages into the gastrointestinal tract and redistribution of muscularis macrophages away from myenteric ganglia. The permanent intraluminal midcerebral artery occlusion model caused changes in gene expression in muscularis macrophages consistent with an altered phenotype. While the size of myenteric ganglia after stroke was not altered, myenteric neurons from post-permanent intraluminal midcerebral artery occlusion mice showed a reduction in neuronal nitric oxide synthase expression, and this response was associated with enhanced intestinal smooth muscle contraction exvivo. Finally, chemical sympathectomy with 6-hydroxydopamine prevented the loss of myenteric neuronal nitric oxide synthase expression and stroke-induced slowed gut transit. Our findings demonstrate that activation of the sympathetic nervous system after stroke is associated with reduced neuronal nitric oxide synthase expression in myenteric neurons, resulting in impaired smooth muscle relaxation and dysregulation of gut transit.

Paxillin participates in the sphingosylphosphorylcholine-induced abnormal contraction of vascular smooth muscle by regulating Rho-kinase activation

BackgroundThe Ca2+-independent contraction of vascular smooth muscle is a leading cause of cardiovascular and cerebrovascular spasms. In the previous study, we demonstrated the involvement of Src family protein tyrosine kinase Fyn and Rho-kinase in the sphingosylphosphorylcholine (SPC)-induced abnormal and Ca2+-independent contraction of vascular smooth muscle, but the specific mechanism has not been completely clarified.MethodsPaxillin knockdown human coronary artery smooth muscle cells (CASMCs) and smooth muscle-specific paxillin knockout mice were generated by using paxillin shRNA and the tamoxifen-inducible Cre-LoxP system, respectively. CASMCs contraction was observed by time-lapse recording. The vessel contractility was measured by using a myography assay. Fyn, Rho-kinase, and myosin light chain activation were assessed by immunoprecipitation and western blotting. The paxillin expression and actin stress fibers were visualized by histological analysis and immunofluorescent staining.ResultsThe SPC-induced abnormal contraction was inhibited in paxillin knockdown CASMCs and arteries of paxillin knockout mice, indicating that paxillin is involved in this abnormal contraction. Further study showed that paxillin knockdown inhibited the SPC-induced Rho-kinase activation without affecting Fyn activation. In addition, paxillin knockdown significantly inhibited the SPC-induced actin stress fiber formation and myosin light chain phosphorylation. These results suggest that paxillin, as an upstream molecule of Rho-kinase, is involved in the SPC-induced abnormal contraction of vascular smooth muscle.ConclusionsThe present study demonstrated that paxillin participates in the SPC-induced abnormal vascular smooth muscle contraction by regulating Rho-kinase activation.Ak4aWCsjd4r4atTQ7PUYn2Video

Esophageal body adaptation to Nissen fundoplication: Increased esophagogastric outflow resistance yields delayed and sustained peristaltic contractions without increased amplitude.

Improvement in lower esophageal sphincter (LES) competency after laparoscopic Nissen fundoplication (LNF) is well established, yet esophageal body physiology data are limited. We aimed to describe the impact of LNF on whole esophagus physiology using standard and novel manometric characteristics. A cohort of patients with an intact fundoplication without herniation and no postoperative dysphagia were selected and underwent esophageal manometry at one-year after surgery. Pre- and post-operative manometry files were reanalyzed using standard and novel manometric characteristics and compared. A total of 95 patients were included in this study. At 16.1 (8.7) months LNF increased LES overall and abdominal length and resting pressure (p < 0.0001). Outflow resistance (IRP) increased [5.8 (3-11) to 11.1 (9-15), p < 0.0001] with a 95th percentile of 20 mmHg in this cohort of dysphagia-free patients. Distal contractile integral (DCI) also increased [1177.0 (667-2139) to 1321.1 (783-2895), p = 0.002], yet contractile amplitude was unchanged (p = 0.158). There were direct correlations between pre- and post-operative DCI [R: 0.727 (0.62-0.81), p < 0.0001] and postoperative DCI and postoperative IRP [R: 0.347 (0.16-0.51), p = 0.0006]. Contractile front velocity [3.5 (3-4) to 3.2 (3-4), p = 0.0013] was slower, while distal latency [6.7 (6-8) to 7.4 (7-9), p < 0.0001], the interval from swallow onset to proximal smooth muscle initiation [4.0 (4-5) to 4.4 (4-5), p = 0.0002], and the interval from swallow onset to point when the peristaltic wave meets the LES [9.4 (8-10) to 10.3 (9-12), p < 0.0001] were longer. Esophageal length [21.9 (19-24) to 23.2 (21-25), p < 0.0001] and transition zone (TZ) length [2.2 (1-3) to 2.5 (1-4), p = 0.004] were longer. Bolus clearance was inversely correlated with TZ length (p = 0.0002) and time from swallow onset to proximal smooth muscle initiation (p < 0.0001). Bolus clearance and UES characteristics were unchanged (p > 0.05). Increased outflow resistance after LNF required an increased DCI. However, this increased contractile vigor was achieved through sustained, not stronger, peristaltic contractions. Increased esophageal length was associated with increased TZ and delayed initiation of smooth muscle contractions.

Hesperetin Relaxes Depolarizing Contraction in Human Umbilical Vein by Inhibiting L-Type Ca2+ Channel.

To study hesperetin-induced vasorelaxation after depolarizing contraction in human umbilical veins (HUVs) to elucidate the role of L-type Ca2+ channel (LTCC) and related signaling pathway. Isometric tension recording was performed in HUV rings pre-contracted with K+. Hesperetin relaxing mechanism was investigated using a LTCC opener (BayK8644) and blockers of cyclic nucleotides and phosphodiesterases (PDEs). Whole-cell patch-clamping in A7r5 cells, a rat vascular smooth muscle cell line, was performed to study the effect of hesperetin on LTCC current. After depolarizing precontraction, hesperetin induced HUV relaxation concentration-dependently and endothelium-independently; 1 mmol/L hesperetin reduced denuded HUV ring tension by 68.7% ± 4.3% compared to matching vehicle, osmolality, and time controls (P<0.0001). Importantly, hesperetin competitively inhibited BayK8644-induced contraction, shifting the half maximal effective concentration of BayK8644 response from 1.08 nmol/L [95% confidence interval (CI) 0.49-2.40] in vehicle control to 11.30 nmol/L (95% CI 5.45-23.41) in hesperetin (P=0.0001). Moreover, hesperetin elicited further vasorelaxation in denuded HUV rings pretreated with inhibitors of soluble guanylyl cyclase, adenylyl cyclase, PDE3, PDE4, and PDE5 (P<0.01), while rings pretreated with PDE1 inhibitors could not be relaxed by hesperetin (P>0.05). However, simultaneously applying inhibitors of soluble guanylyl cyclase and adenylyl cyclase could not inhibit hesperetin's effect (P>0.05). In whole-cell patch-clamping, hesperetin rapidly decreased LTCC current in A7r5 cells to 66.7% ± 5.8% (P=0.0104). Hesperetin diminishes depolarizing contraction of human vascular smooth muscle through inhibition of LTCC, and not cyclic nucleotides nor PDEs. Our evidence supports direct LTCC interaction and provides additional basis for the use of hesperetin and its precursor hesperidin as vasodilators and may lead to future vasodilator drug development as a treatment alternative for cardiovascular diseases.

Abstract A068: Proteomic analysis of keratin 17 positive cells from laser-capture microdissected pancreatic ductal adenocarcinoma

Abstract Introduction. Although patients with pancreatic ductal adenocarcinoma (PDAC) have one of the lowest survival of all carcinomas, there is considerable variation in survival and disease progression, even when accounting for factors such as age and pathologic stage. Our group identified keratin 17 (K17) as a negative prognostic and predictive biomarker in PDAC. Patients with high K17 expression in their tumors have shorter overall survival as well as decreased response to chemotherapeutic agents, including gemcitabine. K17 has also emerged as an established biomarker of the basal molecular subtype of PDAC. In this study, we aim to identify proteins which correlate, directly or indirectly, with K17 expression via a combined approach using immunohistochemistry (IHC) followed by laser capture microdissection (LCM) and mass spectrometry. Methods. We selected 5 PDAC cases of low K17 expression and 5 PDAC cases of high K17 expression from the archival collections from the Stony Brook Cancer Center Biobank. A modified indirect immunoperoxidase approach was performed on sections mounted on LCM-compatible membrane slides. Carcinoma cells that were either K17 positive or negative by IHC were precisely dissected using LMD7 microscope (Leica) from 10-µM thick sections, followed by a modified tissue digestion and protein extraction protocol. Collision-induced fragment mass information and peptide abundance values were obtained by a liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS). Spectral cutoffs were analyzed at both &amp;lt;5% and 1% false discovery rates. The human UniProt dataset was used for data alignment. Results. Mass spectrometry yielded over 1600 detected proteins with approximately 50 of which correlated strongly with K17 expression. Our preliminary results show that keratin 5, a known marker of basal-like PDAC, is positively correlated with keratin 17. Genes whose increased expression contribute to the Moffitt classification of basal molecular subtype of PDAC, such as S100A2, keratin 6A and keratin 7, also showed positive correlation with K17 in our study. The remainder of proteins associated with K17 expression include those involved with cell adhesion, cell-extracellular matrix interaction and smooth muscle contraction pathways, with specific types and classes being identified, pending in-depth analysis. Proteins with inverse relationship to K17 expression included those involved in intracellular organelle trafficking and apoptosis pathways. Conclusion. Our findings demonstrate the utility of mass spectrometry-based spatial proteomic analysis of microdissected cell populations from formalin-fixed, paraffin embedded samples. These methods have the potential to uncover complementary biomarkers to K17 that could enhance prognostic accuracy or serve as therapeutic targets and provide mechanistic insight into the mechanisms through which K17 drives tumor aggression. Citation Format: Ji Dong K. Bai, John D. Haley, Natalia Marchenko, Luisa F. Escobar-Hoyos, Kenneth R. Shroyer. Proteomic analysis of keratin 17 positive cells from laser-capture microdissected pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A068.

The Effects of Tachykinin1 Gene Products on Prepubertal Dabry's Sturgeon (Acipenser dabrynus) Pituitary Hormone Secretion and Gene Expression.

As an ancient and endangered species unique to the Yangtze River in China, the wild population of the Dabry's sturgeon has become scarce. Due to the long time till the first sexual maturity of Dabry's sturgeon, the population of artificially bred Dabry's sturgeon recovered slowly. As a member of the tachykinin family, TAC1 has been reported to have a variety of functions in mammals such as pain control, smooth muscle contraction and reproductive cycle regulation, but the function of Tac1 in fish has been rarely reported. In this study, we synthesized two tac1 gene products, Substance P (SP) and neurokinin A (NKA), and further verified the effect of two tac1 gene products on the secretion of related hormones in the pituitary of Dabry's Sturgeon by intraperitoneal injection and co-culture of primary cells. Expression studies revealed that the newly cloned tac1 were mainly distributed in the hypothalamus and pituitary tissue of the brain. In prepubertal Dabry's sturgeon, this study showed that the two gonadotropins' mRNA levels in pituitary tissue can be significantly increased by SP and NKA through intraperitoneal injection, and the LH protein level in serum was also increased. Further study showed that both NKA and SP could promote the two gonadotropins' mRNA expression in pituitary cells of Dabry's sturgeon. In addition, we explored the optimal dose and time of SP and NKA on pituitary cells is 24 h and over 10 nM. These results, as a whole, suggested that tac1 gene products play an important role in gonadotropin release and gonadal development in prepubertal Dabry's sturgeon.

A computational framework for pharmaco‐mechanical interactions in arterial walls using parallel monolithic domain decomposition methods

AbstractA computational framework is presented to numerically simulate the effects of antihypertensive drugs, in particular calcium channel blockers, on the mechanical response of arterial walls. A stretch‐dependent smooth muscle model by Uhlmann and Balzani is modified to describe the interaction of pharmacological drugs and the inhibition of smooth muscle activation. The coupled deformation‐diffusion problem is then solved using the finite element software FEDDLib and overlapping Schwarz preconditioners from the Trilinos package FROSch. These preconditioners include highly scalable parallel GDSW (generalized Dryja–Smith–Widlund) and RGDSW (reduced GDSW) preconditioners. Simulation results show the expected increase in the lumen diameter of an idealized artery due to the drug‐induced reduction of smooth muscle contraction, as well as a decrease in the rate of arterial contraction in the presence of calcium channel blockers. Strong and weak parallel scalability of the resulting computational implementation are also analyzed.

Blockage of VEGF function by bevacizumab alleviates early-stage cerebrovascular dysfunction and improves cognitive function in a mouse model of Alzheimer’s disease

BackgroundAlzheimer's disease (AD) is a neurodegenerative disorder and the predominant type of dementia worldwide. It is characterized by the progressive and irreversible decline of cognitive functions. In addition to the pathological beta-amyloid (Aβ) deposition, glial activation, and neuronal injury in the postmortem brains of AD patients, increasing evidence suggests that the often overlooked vascular dysfunction is an important early event in AD pathophysiology. Vascular endothelial growth factor (VEGF) plays a critical role in regulating physiological functions and pathological changes in blood vessels, but whether VEGF is involved in the early stage of vascular pathology in AD remains unclear.MethodsWe used an antiangiogenic agent for clinical cancer treatment, the humanized monoclonal anti-VEGF antibody bevacizumab, to block VEGF binding to its receptors in the 5×FAD mouse model at an early age. After treatment, memory performance was evaluated by a novel object recognition test, and cerebral vascular permeability and perfusion were examined by an Evans blue assay and blood flow scanning imaging analysis. Immunofluorescence staining was used to measure glial activation and Aβ deposits. VEGF and its receptors were analyzed by enzyme-linked immunosorbent assay and immunoblotting. RNA sequencing was performed to elucidate bevacizumab-associated transcriptional signatures in the hippocampus of 5×FAD mice.ResultsBevacizumab treatment administered from 4 months of age dramatically improved cerebrovascular functions, reduced glial activation, and restored long-term memory in both sexes of 5×FAD mice. Notably, a sex-specific change in different VEGF receptors was identified in the cortex and hippocampus of 5×FAD mice. Soluble VEGFR1 was decreased in female mice, while full-length VEGFR2 was increased in male mice. Bevacizumab treatment reversed the altered expression of receptors to be comparable to the level in the wild-type mice. Gene Set Enrichment Analysis of transcriptomic changes revealed that bevacizumab effectively reversed the changes in the gene sets associated with blood–brain barrier integrity and vascular smooth muscle contraction in 5×FAD mice.ConclusionsOur study demonstrated the mechanistic roles of VEGF at the early stage of amyloidopathy and the protective effects of bevacizumab on cerebrovascular function and memory performance in 5×FAD mice. These findings also suggest the therapeutic potential of bevacizumab for the early intervention of AD.

The Bronchodilator Potential of Astragalus sarcocolla: An in vitro Experiment.

To evaluate the bronchodilatory mechanism of Astragalus sarcocolla (ASE) extract on tracheal smooth muscles of rabbits. In-vitro experimental study. Place and Duration of the Study: The animal house of CMH Lahore Medical College, Lahore, and Institute of Dentistry, NUMS, from October 2022 to May 2023. Six rabbits were randomly divided into four groups. After euthanising the rabbit, the trachea was carefully dissected out and stabilised in Kreb's Henseleit solution for 30 minutes and then, stimulated by acetylcholine (Ach) 1µm, under mimicked physiological conditions. Group I served as the control group with tracheal smooth muscles stabilised with 1g tension. In Group II (positive control), tracheal smooth muscles were stimulated by potassium chloride (KCl) (80 mM and 25 mM, respectively) to get maximum tracheal smooth muscle contractions. Later, the tissue was exposed to theophylline with three molar concentrations 0.2, 0.4, 0.6, and 0.8 mM, and cumulative dose response curves were formed. In Group III (ASE group), tracheal smooth muscles were stimulated by KCl (80 mM and 25 mM) and was exposed to increasing concentration of ASE. In group IV, tissue was stimulated by KCl (25 mM) and glibenclamide (3 µM), later exposed to increasing concentration of ASE to confirm the bronchodilatory mechanism. The change in isometric contraction of the tissue was recorded using the force displacement transducer connected to a PowerLab data acquisition system. Concentration response curves were drawn, and median effective concentrations (EC50 values) and percentage inhibition were calculated. Non-linear regression was applied for the analysis of the concentration-response curves. ASE inhibited the KCl-induced low potassium (25 mM) contractions (EC50 = 0.38 mg/ml, 95% CI: 0.04 - 0.38, n = 6). It only partially inhibited the high potassium-induced contractions in tracheal smooth muscles. Pretreatment with glibenclamide showed a rightward shift of the dose-response curve. Theophylline and ASE significantly reduced the low K+ induced smooth muscle contractions in comparison to the control group (p <0.001, each). Astragalus sarcocolla extract produced bronchodilator effects through the activation of ATP sensitive potassium channels in isolated rabbit trachea. Astragalus sarcocolla, Bronchodilators, ATP-sensitive potassium channels, Effective concentration 50, Concentration response curves.

Desmopressin nasal spray inhibiting parasympathetic function on isolated tracheal smooth muscle.

Objectives: Nocturia with or without asthma is one of the aging diseases. Desmopressin has been used as a nasal spray for patients who are suffering from nocturia. This study determined the effects of desmopressin on isolated tracheal smooth muscle in vitro. Methods: We evaluated desmopressin's efficiency on isolated rat tracheal smooth muscle. Desmopressin was evaluated for the following effects on tracheal smooth muscle: (1) effect on resting tension; (2) effect on contraction brought on by parasympathetic mimetic 10-6 M methacholine; and (3) effect on electrically produced tracheal smooth muscle contractions. Results: As the concentration grew, desmopressin by itself had no impact on the trachea's baseline tension. Addition of desmopressin at doses of 10-5 M or above elicited a significant relaxation response to 10-6 M methacholine-induced contraction. Desmopressin could also inhibit spike contraction of the trachea induced by electrical field. Conclusion: According to this study, desmopressin at high quantities may prevent the trachea's parasympathetic activity. Due to its ability to block parasympathetic activity and lessen the contraction of the tracheal smooth muscle brought on by methacholine, Desmopressin nasal spray might help nocturia sufferers experience fewer asthma attacks.

Difference in Contractile Mechanisms between the Early and Sustained Components of Ionomycin-Induced Contraction in Rat Caudal Arterial Smooth Muscle.

We previously reported that the sustained component of contraction induced by depolarizing stimulation by high K+ concentration in rat caudal arterial smooth muscle involves a Ca2+-induced Ca2+ sensitization mechanism whereby Ca2+ entry through voltage-gated Ca2+ channels activates proline-rich tyrosine kinase 2 (Pyk2), leading to activation of RhoA/Rho-associated kinase (ROCK). In the present study, we investigated a potential role for Pyk2-mediated RhoA/ROCK activation in contraction mediated by elevation of cytosolic free Ca2+ concentration ([Ca2+]i) induced by a Ca2+ ionophore, ionomycin, rather than by depolarizing stimulation. Ionomycin (60 µM) induced slow and sustained contraction of rat caudal arterial smooth muscle due to influx of Ca2+. Pre-treatment with a myosin light chain kinase (MLCK) inhibitor, ML-9 (30 µM), inhibited both the early phase (4 min) and the sustained phase (30 min) of ionomycin-induced contraction. On the other hand, a ROCK inhibitor, HA-1077 (3 µM), and Pyk2 inhibitors, sodium salicylate (10 mM) and PF-431396 (3 µM), suppressed only the sustained phase of ionomycin-induced contraction. A calmodulin (CaM) inhibitor, W-7 (150 µM), but not W-5 (150 µM), suppressed the early phase of contraction. Early or sustained increase of ionomycin-induced 20 kDa light chain of myosin (LC20) phosphorylation was inhibited by each inhibitor in a manner similar to the attenuation of contraction. These results indicate that the early phase of ionomycin-induced contraction is mediated by MLCK activation by [Ca2+]i elevation, whereas the sustained phase of ionomycin-induced contraction involves RhoA/ROCK activation and inhibition of myosin light chain phosphatase (MLCP) through CaM-independent Pyk2 activation by [Ca2+]i elevation.

Chloroquine and cinchonine affect rat vascular smooth muscle tonus through calcium channels - in silico and in vitro approaches.

In the present study, two structurally similar alkaloids from trees of Cinchona genus, chloroquine and cinchonine, were examined for their vasorelaxant effects in amodel of phenylephrine-induced smooth muscle contractions. Potential mechanisms of action associated with endothelial vasorelaxant compounds, voltage-gated Ca2+ channels (LTCCs), and inositol triphosphate receptors were examined in isolated rat aortic rings. Also, an in silico approach was used to predict the activity of the two test compounds. Experimental results revealed that both chloroquine and cinchonine significantly decrease phenylephrine-induced smooth muscle contractions, although to adifferent extent. Evaluated mechanisms of action indicate that endothelium is not involved in the vasorelaxant action of the two tested alkaloids. On the other hand, voltage-gated Ca2+ channels were found to be the dominant way of action associated with the vasorelaxant action of chloroquine and cinchonine. Finally, IP3R is found to have only asmall impact on the observed activity of the tested compounds. Molecular docking studies predicted that chloroquine possesses asignificant activity toward asuitable model of LTCCs, while cinchonine does not. The results of the present study point to the fact that great caution should be paid while administering chloroquine to vulnerable patients, especially those with cardiovascular disorders (Tab. 3, Fig. 3, Ref. 28).

DREADD agonist compound 21 causes acute diuresis in wild-type mice.

The targeted activation or inhibition of specific cell populations using chemogenetics allows the precise dissection of cellular signaling and function. Designer receptors exclusively activated by designer drugs (DREADDs) is a chemogenetic platform initially developed by mutating human muscarinic receptors to be unresponsive to endogenous acetylcholine but exclusively activated by an "inert" designer drug. Compound 21 (C21) is a new and potent DREADD agonist; however, radioligand assays from a recent report indicated its ability to bind to endogenous G protein-coupled receptors (GPCRs), including muscarinic M1-3 receptors. Whether this binding causes off-target effects is unclear. Renal innervation is important for the regulation of renal function, and the advent of chemogenetic tools provides significant opportunities for the mechanistic understanding of renal innervation and function. GPCRs such as adrenergic and muscarinic receptors play a role in renal function; thus, a careful pharmacological characterization of C21 in renal function is a prerequisite for this approach. Unexpectedly, an infusion of 1.0mg/kg C21 in anesthetized mice caused an ∼4-fold increase in urine output and correspondingly increased the glomerular filtration rate (GFR), suggesting a C21-mediated acute diuretic effect. This acute diuresis effect was further confirmed in awake mice using voiding spot assays. The exact molecular mechanism for C21-mediated diuresis is unclear; however, we demonstrated by in vitro myography that C21 can effectively inhibit bladder smooth muscle contraction by antagonizing M3 receptors at the micromolar level, causing increased voiding size in vivo. In summary, C21 functions as a GPCR antagonist and has significant dose-dependent off-target effects in the renal system.

Origin and Development of Interstitial Cells of Cajal.

The digestive tract is a series of organs with specific functions and specialized anatomy. Each organ is organized similarly with concentric layers of epithelial, connective, smooth muscle, and neural tissues. Interstitial cells of Cajal (ICC) are distributed in smooth muscle layers and contribute to the organization of repetitive and rhythmic smooth muscle contractions. Understanding ICC development is critical to understanding gastrointestinal motility patterns. Experiments determining ICC origin and development in mice, chicken, and humans are described, as well as what is known in the zebrafish. At least six types of ICC in the digestive tract have been described and ICC heterogeneity in adult tissues is reviewed. Factors required for ICC development and for maintenance of ICC subclasses are described. This review is suitable for those new to ICC development and physiology, especially those focused on using zebrafish and other model systems.