Vasoactive Intestinal Peptide Receptor 2 Research Articles

Blockade of vasoactive intestinal peptide receptor 2 (VIPR2) signaling suppresses cyclin D1-dependent cell-cycle progression in MCF-7 cells

Vasoactive intestinal peptide (VIP) receptor 2 (VIPR2) is a G protein-coupled receptor that binds to Gαs, Gαi, and Gαq proteins to regulate various downstream signaling molecules, such as protein kinase A (PKA), phosphatidylinositol 3-kinase (PI3K), and phospholipase C. In this study, we examined the role of VIPR2 in cell cycle progression. KS-133, a newly developed VIPR2-selective antagonist peptide, attenuated VIP-induced cell proliferation in MCF-7 cells. The percentage of cells in the S-M phase was decreased in MCF-7 cells treated with KS-133. KS-133 in the presence of VIP decreased the phosphorylation of extracellular signal-regulated kinase (ERK), AKT, and glycogen synthase kinase-3β (GSK3β), resulting in a decrease in cyclin D1 levels. In MCF-7 cells stably-expressing VIPR2, KS-133 decreased PI3K activity and cAMP levels. Treatment with the ERK-specific kinase (MEK) inhibitor U0126 and the class I PI3K inhibitor ZSTK474 decreased the percentage of cells in the S phase. KS-133 reduced the percentage of cells in the S phase more than treatment with U0126 or ZSTK474 alone and did not affect the effect of the mixture of these inhibitors. Our findings suggest that VIPR2 signaling regulates cyclin D1 levels through the cAMP/PKA/ERK and PI3K/AKT/GSK3β pathways, and mediates the G1/S transition to control cell proliferation.

Development of a New Enzyme-Linked Immunosorbent Assay (ELISA) for Measuring the Content of PACAP in Mammalian Tissue and Plasma.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a naturally occurring neuropeptide found in both the central and peripheral nervous systems of vertebrates. Recent studies have revealed the presence of PACAP and its corresponding receptors, namely, the pituitary adenylate cyclase-activating polypeptide type I receptor (PAC1R), vasoactive intestinal peptide receptor 1 (VIPR1), and vasoactive intestinal peptide receptor 2 (VIPR2), in various structures implicated in migraine pathophysiology, including sensory trigeminal neurons. Human studies have demonstrated that when infused, PACAP can cause dilation of cranial vessels and result in delayed migraine-like attacks. In light of this, we present a novel ELISA assay that has been validated for quantifying PACAP in tissue extracts and human plasma. Using two well characterized antibodies specifically targeting PACAP, we successfully developed a sandwich ELISA assay, capable of detecting and accurately quantifying PACAP without any cross-reactivity to closely related peptides. The quantification range was between 5.2 pmol/L and 400 pmol/L. The recovery in plasma ranged from 98.2% to 100%. The increasing evidence pointing to the crucial role of PACAP in migraine pathophysiology necessitates the availability of tools capable of detecting changes in the circulatory levels of PACAP and its potential application as a reliable biomarker.

The VIPR2-selective antagonist KS-133 changes macrophage polarization and exerts potent anti-tumor effects as a single agent and in combination with an anti-PD-1 antibody.

We have previously demonstrated that KS-133 is a specific and potent antagonist of vasoactive intestinal peptide receptor 2 (VIPR2). We have also shown that vasoactive intestinal peptide-VIPR2 signaling affects the polarity and activation of tumor-associated macrophages, which is another strategy for cancer immunotherapy apart from the activation of effector T cells. In this study, we aimed to examine whether the selective blockade of VIPR2 by KS-133 changes the polarization of macrophages and induces anti-tumor effects. In the presence of KS-133, genetic markers indicative of tumor-aggressive M1-type macrophages were upregulated, and conversely, those of tumor-supportive M2-type macrophages were downregulated. Daily subcutaneous administration of KS-133 tended to suppress the growth of CT26 tumors (murine colorectal cancer-derived cells) implanted subcutaneously in Balb/c mice. To improve the pharmacological efficacy and reduce the number of doses, we examined a nanoformulation of KS-133 using the US Food and Drug Administration-approved pharmaceutical additive surfactant Cremophor® EL. KS-133 nanoparticles (NPs) were approximately 15 nm in size and stable at 4°C after preparation. Meanwhile, KS-133 was gradually released from the NPs as the temperature was increased. Subcutaneous administration of KS-133 NPs once every 3 days had stronger anti-tumor effects than daily subcutaneous administration of KS-133. Furthermore, KS-133 NPs significantly enhanced the pharmacological efficacy of an immune checkpoint-inhibiting anti-PD-1 antibody. A pharmacokinetic study suggested that the enhancement of anti-tumor activity was associated with improvement of the pharmacokinetic profile of KS-133 upon nanoformulation. Our data have revealed that specific blockade of VIPR2 by KS-133 has therapeutic potential for cancer both alone and in combination with immune checkpoint inhibitors.

Development of the vasoactive intestinal peptide receptor 2 (VIPR2) antagonist peptide for the treatment of schizophrenia

Schizophrenia affects approximately 24 million people worldwide. Existing medications for the treatment of schizophrenia work primarily by improving positive symptoms such as agitation, hallucinations, delusions, and aggression. They possess common mechanism of action (MOA), blocking to neurotransmitter receptors such as dopamine, serotonin, and adrenaline receptors. Although multiple agents are available for the treatment of schizophrenia, the majority do not address negative symptoms or cognitive dysfunction. In other cases, patients have drug-related adverse effects. The vasoactive intestinal peptide receptor 2 (VIPR2, also known as VPAC2 receptor) might be an attractive drug target for the treatment of schizophrenia because both clinical and preclinical studies have demonstrated a strong link between high expression/overactivation of VIPR2 and schizophrenia. Despite these backgrounds, the proof-of-concept of VIPR2 inhibitors has not been examined clinically. A reason might be that VIPR2 belongs to class-B GPCRs, and the discovery of small-molecule drugs against class-B GPCRs is generally difficult. We have developed a bicyclic peptide KS-133, which shows VIPR2 antagonist activity and suppresses cognitive decline in a mouse model relevant to schizophrenia. KS-133 has a different MOA from current therapeutic drugs and exhibits high selectivity for VIPR2 and potent inhibitory activity against a single-target molecule. Therefore, it may contribute to both the development of a novel drug candidate for the treatment of psychiatric disorders such as schizophrenia and acceleration of basic studies on VIPR2.

Vasoactive intestinal peptide receptor 2 signaling promotes breast cancer cell proliferation by enhancing the ERK pathway

Vasoactive intestinal peptide (VIP) receptor 2 (VIPR2) is a class B G protein-coupled receptor with the neuropeptide VIP as a ligand. Increased VIPR2 mRNA expression and/or VIPR2 gene copy number has been documented in several cancers including breast carcinoma. However, the pathophysiological role of increased VIPR2 in the proliferation of breast cancer cells remains largely unknown. In this study, we found that VIPR2 overexpression in MCF-7 and MDA-MB-231 cells, human breast cancer cell lines, promoted cell proliferation. Increased VIPR2 also exacerbated intraperitoneal proliferation of breast cancer MDA-MB-231 cells in a tumor nude mouse model in vivo. Treatment with KS-133, a VIPR2-selective antagonist peptide, significantly inhibited VIP-induced cell proliferation in VIPR2-overexpressing MCF-7 and MDA-MB-231 cells. Overexpressed VIPR2 caused increases in the levels of cAMP and phosphorylated extracellular signal-regulated kinase (ERK), which involves a VIPR2 signaling pathway through Gs protein. Additionally, phosphorylation of vasodilator-stimulated phosphoprotein (Ser157) and cAMP response element binding protein (Ser133) in VIPR2-overexpressing MCF-7 cells was greater than that in control cells, suggesting the increased PKA activity. Moreover, an inhibitor of mitogen-activated protein kinase kinase, U0126, attenuated tumor proliferation in exogenous VIPR2-expressing MCF-7 and MDA-MB-231 cells at the same level as observed in EGFP-expressing cells treated with U0126. Together, these findings suggest that VIPR2 controls breast tumor growth by regulating the cAMP/PKA/ERK signaling pathway, and the excessive expression of VIPR2 may lead to an exacerbation of breast carcinoma.

AlphaFold version 2.0 elucidates the binding mechanism between VIPR2 and KS-133, and reveals an S–S bond (Cys25−Cys192) formation of functional significance for VIPR2

The vasoactive intestinal peptide receptor 2 (VIPR2) has attracted attention as a drug target for the treatment of mental disorders, cancer, and immune diseases. In 2021, we identified the peptide KS-133 as a VIPR2-selective antagonist. In this study, we aimed to elucidate the binding mechanism between VIPR2 and KS-133. To this end, VIPR2/KS-133 and VIPR2/vasoactive intestinal peptide (VIP) complex models were constructed through AlphaFold version 2.0 and molecular dynamic simulations. Our models revealed that: (i) both KS-133 and VIP have helical structures, (ii) the interaction residues on VIPR2 for both peptides are similar, and (iii) the orientation of their helices upon their binding to VIPR2 are different by ∼45°. Interestingly, in the process of constructing the aforementioned models, an S–S bond formation between Cys25 and Cys192 of the human VIPR2 was identified. Although these two Cys residues are highly conserved among species (i.e., corresponding to Cys24 and Cys191 in the mouse), no previous reports regarding this S–S bond formation exist. In order to clarify the potential role of this S–S bond in the VIPR2 has functional consequences, a cell line expressing the mouse VIPR2(Cys24Ala, Cys191Ala) was generated. During the VIP stimulation of this cell line, the phosphorylation of AKT (a downstream signal marker of VIPR2) was found to be significantly attenuated, thereby suggesting that the S–S bond has a functional significance for VIPR2. Our study not only elucidates the VIPR2-binding mechanism of KS-133 for the first time, but also provides new insights into the structural biology of VIPR2.

Generation of KS-133 as a Novel Bicyclic Peptide with a Potent and Selective VIPR2 Antagonist Activity that Counteracts Cognitive Decline in a Mouse Model of Psychiatric Disorders.

Worldwide, more than 20 million people suffer from schizophrenia, but effective and definitive new therapeutic drugs/treatments have not been established. Vasoactive intestinal peptide receptor 2 (VIPR2) might be an attractive drug target for the treatment of schizophrenia because both preclinical and clinical studies have demonstrated a strong link between high expression/overactivation of VIPR2 and schizophrenia. Nevertheless, VIPR2-targeting drugs are not yet available. VIPR2 is a class-B G protein-coupled receptor that possesses high structural homology to its subtypes, vasoactive intestinal peptide receptor 1 (VIPR1) and pituitary adenylate cyclase-activating polypeptide type-1 receptor (PAC1). These biological and structural properties have made it difficult to discover small molecule drugs against VIPR2. In 2018, cyclic peptide VIpep-3, a VIPR2-selective antagonist, was reported. The aim of this study was to generate a VIpep-3 derivative for in vivo experiments. After amino acid substitution and structure optimization, we successfully generated KS-133 with 1) a VIPR2-selective and potent antagonistic activity, 2) at least 24 h of stability in plasma, and 3) in vivo pharmacological efficacies in a mouse model of psychiatric disorders through early postnatal activation of VIPR2. To the best of our knowledge, this is the first report of a VIPR2-selective antagonistic peptide that counteracts cognitive decline, a central feature of schizophrenia. KS-133 may contribute to studies and development of novel schizophrenia therapeutic drugs that target VIPR2.

Tamoxifen-inducible cardiac-specific Cre transgenic mouse using VIPR2 intron

Genetically engineered mouse models through gene deletion are useful tools for analyzing gene function. To delete a gene in a certain tissue temporally, tissue-specific and tamoxifen-inducible Cre transgenic mice are generally used. Here, we generated transgenic mouse with cardiac-specific expression of Cre recombinase fused to a mutant estrogen ligand-binding domain (ERT2) on both N-terminal and C-terminal under the regulatory region of human vasoactive intestinal peptide receptor 2 (VIPR2) intron and Hsp68 promoter (VIPR2-ERT2CreERT2). In VIPR2-ERT2CreERT2 transgenic mice, mRNA for Cre gene was highly expressed in the heart. To further reveal heart-specific Cre expression, VIPR2-ERT2CreERT2 mice mated with ROSA26-lacZ reporter mice were examined by X-gal staining. Results of X-gal staining revealed that Cre-dependent recombination occurred only in the heart after treatment with tamoxifen. Taken together, these results demonstrate that VIPR2-ERT2CreERT2 transgenic mouse is a useful model to unveil a specific gene function in the heart.

In vivo and in vitro effects of Salsola collina on gastrointestinal motility in rats.

Objective(s): Salsola collina is widely distributed along the Bohai coast and consumed as an edible plant by native residents. We have found surprisingly that S. collina extracts promoted gastrointestinal motility in mice previously. In the present study, effects of S. collina on gastrointestinal motility in rats and its underlying mechanism were explored.Materials and Methods: In vivo, different fraction extracts from S. collina were prepared and the effects on gastric emptying and small intestinal propulsion in normal rats were measured. Plasma ghrelin (GRL), motilin (MTL), gastrin (GAS) and vasoactive intestinal peptide (VIP) and expressions of GRL receptor (GHSR), MTL receptor (MTLR), VIP receptor 2 (VIPR2) in the duodenum were also detected. In vitro, gastric antrum strips were prepared and activities of different extracts on gastric smooth muscle contractions were evaluated.Results:Results showed that the ethyl acetate extract (EAE) was the most effective fraction to promote gastric emptying and intestinal propulsion, showing a dose-dependent manner. EAE increased plasma GRL and GAS, elevated GHSR expression and restrained VIPR2 expression in the duodenum. In vitro, EAE promoted contraction of normal gastric antrum strips as well as relaxed strips induced by atropine.Conclusion:These data indicate that EAE has a significant prokinetic activity via a mechanism that mainly involves in modulating plasma GRL and GAS, expressions of GSHR and VIPR2 in the duodenum and activating M-cholinergic receptor. Our study provides a pharmacological basis for the use of S. collina extract in treating gastrointestinal motility disorders.

The neuropeptide VIP confers anticipatory mucosal immunity by regulating ILC3 activity.

Group 3 innate lymphoid cell (ILC3)-mediated production of the cytokine interleukin-22 (IL-22) is critical for the maintenance of immune homeostasis in the gastrointestinal tract. Here, we find that the function of ILC3s is not constant across the day, but instead oscillates between active phases and resting phases. Coordinate responsiveness of ILC3s in the intestine depended on the food-induced expression of the neuropeptide vasoactive intestinal peptide (VIP). Intestinal ILC3s had high expression of the G protein-coupled receptor vasoactive intestinal peptide receptor 2 (VIPR2), and activation by VIP markedly enhanced the production of IL-22 and the barrier function of the epithelium. Conversely, deficiency in signaling through VIPR2 led to impaired production of IL-22 by ILC3s and increased susceptibility to inflammation-induced gut injury. Thus, intrinsic cellular rhythms acted in synergy with the cyclic patterns of food intake to drive the production of IL-22 and synchronize protection of the intestinal epithelium through a VIP-VIPR2 pathway in ILC3s.

The myopia susceptibility locus vasoactive intestinal peptide receptor 2 (VIPR2) contains variants with opposite effects

Myopia is the commonest eye disorder in the world. High myopes are predisposed to ocular pathologies. The vasoactive intestinal peptide receptor 2 (VIPR2) gene was identified as a myopia susceptibility locus by our group and another group. We continued to fine-map this locus. A case-control study was performed in 4 sequential stages with a total of 941 highly myopic subjects and 846 control subjects, all unrelated Chinese. Stage 1 experimentally genotyped 64.4% of the entire cohort for 152 single-nucleotide polymorphisms (SNPs) and Stage 2 the remaining subjects for 21 SNPs. Stage 3 combined the genotypes for 21 SNPs for the entire cohort, and identified one group of high-risk haplotypes and one group of protective haplotypes significantly associated with high myopia. Stage 4 imputed genotypes for variants in the VIPR2 region and identified two independent groups of variants: one group with high-risk minor alleles and another with protective minor alleles. Variants within each group were generally in strong linkage disequilibrium among themselves while high-risk variants were in linkage equilibrium with protective variants. Therefore, the VIPR2 locus seems to contain variants with opposite effects. This is the first study that has examined the genetic architecture of a myopia susceptibility locus in detail.

A 3' UTR SNP rs885863, a cis-eQTL for the circadian gene VIPR2 and lincRNA 689, is associated with opioid addiction.

There is a reciprocal relationship between the circadian and the reward systems. Polymorphisms in several circadian rhythm-related (clock) genes were associated with drug addiction. This study aims to search for associations between 895 variants in 39 circadian rhythm-related genes and opioid addiction (OUD). Genotyping was performed with the Smokescreen® array. Ancestry was verified by principal/MDS component analysis and the sample was limited to European Americans (EA) (OUD; n = 435, controls; n = 138). Nominally significant associations (p < 0.01) were detected for several variants in genes encoding vasoactive intestinal peptide receptor 2 (VIPR2), period circadian regulator 2 (PER2), casein kinase 1 epsilon (CSNK1E), and activator of transcription and developmental regulator (AUTS2), but no signal survived correction for multiple testing. There was intriguing association signal for the untranslated region (3’ UTR) variant rs885863 in VIPR2, (p = .0065; OR = 0.51; 95% CI 0.31–0.51). The result was corroborated in an independent EA OUD sample (n = 398, p = 0.0036; for the combined samples). Notably, this SNP is an expression quantitative trait locus (cis-eQTL) for VIPR2 and a long intergenic non-coding RNA, lincRNA 689, in a tissue-specific manner, based on the Genotype-Tissue Expression (GTEx) project. Vasoactive intestinal peptide (VIP) is an important peptide of light-activated suprachiasmatic nucleus cells. It regulates diverse physiological processes including circadian rhythms, learning and memory, and stress response. This is the first report of an association of a VIPR2 variant and OUD. Additionally, analysis of combinations of single nucleotide polymorphisms (SNPs) genotypes revealed an association of PER2 SNP rs80136044, and SNP rs4128839, located 41.6 kb downstream of neuropeptide Y receptor type 1 gene, NPY1R (p = 3.4 × 10−6, OR = 11.4, 95% CI 2.7–48.2). The study provides preliminary insight into the relationship between genetic variants in circadian rhythm genes and long non-coding RNA (lncRNAs) in their vicinity, and opioid addiction.

A Synthetic Agonist to Vasoactive Intestinal Peptide Receptor-2 Induces Regulatory T Cell Neuroprotective Activities in Models of Parkinson's Disease.

A paradigm shift has emerged in Parkinson’s disease (PD) highlighting the prominent role of CD4+ Tregs in pathogenesis and treatment. Bench to bedside research, conducted by others and our own laboratories, advanced a neuroprotective role for Tregs making pharmacologic transformation of immediate need. Herein, a vasoactive intestinal peptide receptor-2 (VIPR2) peptide agonist, LBT-3627, was developed as a neuroprotectant for PD-associated dopaminergic neurodegeneration. Employing both 6-hydroxydopamine (6-OHDA) and α-synuclein (α-Syn) overexpression models in rats, the sequential administration of LBT-3627 increased Treg activity without altering cell numbers both in naïve animals and during progressive nigrostriatal degeneration. LBT-3627 administration was linked to reductions of inflammatory microglia, increased survival of dopaminergic neurons, and improved striatal densities. While α-Syn overexpression resulted in reduced Treg activity, LBT-3627 rescued these functional deficits. This occurred in a dose-dependent manner closely mimicking neuroprotection. Taken together, these data provide the basis for the use of VIPR2 agonists as potent therapeutic immune modulating agents to restore Treg activity, attenuate neuroinflammation, and interdict dopaminergic neurodegeneration in PD. The data underscore an important role of immunity in PD pathogenesis.

Dosage sensitivity intolerance of VIPR2 microduplication is disease causative to manifest schizophrenia-like phenotypes in a novel BAC transgenic mouse model.

Recent genome-wide association studies (GWAS) have identified copy number variations (CNVs) at chromosomal locus 7q36.3 that significantly contribute to the risk of schizophrenia, with all of the microduplications occurring within a single gene: vasoactive intestinal peptide receptor 2 (VIPR2). To confirm disease causality and translate such a genetic vulnerability into mechanistic and pathophysiological insights, we have developed a series of conditional VIPR2 bacterial artificial chromosome (BAC) transgenic mouse models of VIPR2 CNV. VIPR2 CNV mouse model recapitulates gene expression and signaling deficits seen in human CNV carriers. VIPR2 microduplication in mice elicits prominent dorsal striatal dopamine dysfunction, cognitive, sensorimotor gating, and social behavioral deficits preceded by an increase of striatal cAMP/PKA signaling and the disrupted early postnatal striatal development. Genetic removal of VIPR2 transgene expression via crossing with Drd1a-Cre BAC transgenic mice rescued the dopamine D2 receptor abnormality and multiple behavioral deficits, implicating a pathogenic role of VIPR2 overexpression in dopaminoceptive neurons. Thus, our results provide further evidence to support the GWAS studies that the dosage sensitivity intolerance of VIPR2 is disease causative to manifest schizophrenia-like dopamine, cognitive, and social behavioral deficits in mice. The conditional BAC transgenesis offers a novel strategy to model CNVs with a gain-of -copies and facilitate the genetic dissection of when/where/how the genetic vulnerabilities affect development, structure, and function of neural circuits. Our findings have important implications for therapeutic development, and the etiology-relevant mouse model provides a useful preclinical platform for drug discovery.

Predictive Genes for the Prognosis of Central Serous Chorioretinopathy

To investigate potential genetic prognostic factors associated with spontaneous resolution of serous retinal detachment (SRD) and development of choroidal neovascularization (CNV) in central serous chorioretinopathy (CSC). Retrospective analysis of a case series. One hundred ninety-six eyes from 196 patients with active CSC. We retrospectively reviewed medical records and determined the presence or absence of SRD using OCT imaging. The duration until the spontaneous SRD resolution was analyzed using the Kaplan-Meier method, and associations between the duration to spontaneous resolution and Complement factor H (CFH) I62V, Age-Related Maculopathy Susceptibility 2 (ARMS2) A69S, or Vasoactive Intestinal Peptide Receptor 2 (VIPR2) rs3793217 genotypes were evaluated, followed by the assessment of their associations with CNV that developed secondary to CSC. Genetic associations of CFH rs800292, ARMS2 rs10490924, and VIPR2 rs3793217 genotypes with the duration to spontaneous resolution of SRD and development of CNV during follow-up of CSC. In 105 of the 196 study participants, we revealed spontaneous SRD resolution in their eyes during follow-up evaluation. Sixty-eight eyes received treatment, and 23 eyes dropped out before spontaneous SRD resolution. Among the 3 genetic polymorphisms assessed herein, only the CFH I62V genotype was predictive of spontaneous SRD resolution among its genotypes (P= 0.017); the average durations for the spontaneous SRD resolution for the individuals with AA, AG, and GG genotype were 126.6±115.5 days, 157.7±243.1 days, and 242.7±198.0 days, respectively, indicating that the G allele was associated with significantly longer persistent SRD (P= 0.035). Among the total number of eyes of all participants, 14 demonstrated CNV during follow-up evaluation. The CFH I62V G and ARMS2 A69S T alleles were associated significantly with CNV development (P= 0.0023 and P= 0.019, respectively), whereas the VIPR2 rs3793217 genotype was not. The CFH I62V and ARMS2 A69S genotypes can predict the prognosis of CSC. Knowledge of the genetic status may help physicians determine the need for early treatment and possibly prevent subsequent CNV development. Further prospective studies are needed to confirm the observed genotype-phenotype relationship.

Manganese-Enhanced Magnetic Resonance Imaging for Detection of Vasoactive Intestinal Peptide Receptor 2 Agonist Therapy in a Model of Parkinson's Disease.

Neuroprotective immunity is defined by transformation of T-cell polarity for therapeutic gain. For neurodegenerative disorders and specifically for Parkinson's disease (PD), granulocyte-macrophage colony stimulating factor or vasoactive intestinal peptide receptor 2 (VIPR2) agonists elicit robust anti-inflammatory microglial responses leading to neuronal sparing in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-intoxicated mice. While neurotherapeutic potential was demonstrated for PD, there remain inherent limitations in translating these inventions from the laboratory to patients. One obstacle in translating such novel neurotherapeutics centers on the availability of suitable noninvasive methods to track disease progression and therapeutic efficacy. To this end, we developed manganese-enhanced magnetic resonance imaging (MEMRI) assays as a way to track a linkage between glial activation and VIPR2 agonist (LBT-3627)-induced neuroprotective immunity for MPTP-induced nigrostriatal degeneration. Notably, LBT-3627-treated, MPTP-intoxicated mice show reduced MEMRI brain signal intensities. These changes paralleled reduced astrogliosis and resulted in sparing of nigral tyrosine hydroxylase neurons. Most importantly, the data suggest that MEMRI can be developed as a biomarker tool to monitor neurotherapeutic responses that are relevant to common neurodegenerative disorders used to improve disease outcomes.

Attenuation of stress-induced gastrointestinal motility disorder by gentiopicroside, from Gentiana macrophylla Pall

AimThe current study was designed to explore the mechanism of the prokinetic activity of Gentiopicroside (Ge), from Gentiana macrophylla Pall which is widely used to strengthen gastric motility in clinic. MethodsGastrointestinal motility disorder rats were induced by stress stimulation and the rats were treated with Ge. The functions of gastric emptying and intestinal propelling were measured after blood was obtained to assay the levels of plasmatic motilin (MTL), vasoactive intestinal peptide (VIP), somatostatin (SST), gastrin (GAS), neurotensin (NT) and substance of P (SP). The expressions of MTL receptor (MTLR), VIP receptor 2 (VIPR2) and SST receptor 2 (SSTR2) were measured also. In addition, an isolated guinea pig ileum was applied to evaluate the influences of Ge on M-R, H1-R, 5-HT4-R and D-R in vitro. ResultsGe increased gastric emptying and intestinal propelling obviously. It also decreased the level of SST and increased GAS in plasma significantly. Moreover, it promoted the expressions of MTLR in gastric antrum, duodenum, jejunum and ileum, and restrained the expression of VIPR2 in duodenum. Piboserod and loratadine had no obvious restrain to Ge′ exciting ileum effect and Ge also didn't affect dopamine paralyzing ileum. However, Ge failed to improve the hypofunction of guinea pigs ileums pre-treated with atropine sulfate. ConclusionThe mechanisms of Ge′ prokinetic effect were associated with modulating the levels of SST and GAS in plasma, raising the expressions of MTLR in gastric antrum, duodenum, ileum and jejunum, reducing the expression of VIPR2 in duodenum and activating M-R.

A competitive PCR assay confirms the association of a copy number variation in the VIPR2 gene with schizophrenia in Han Chinese

Evidence from genetic studies has revealed that genome-wide rare copy number variations (CNVs) are risk factors for neurodevelopmental disorders and this evidence has given rise to a new understanding of disease etiology, including that of schizophrenia (SCZ). Recent studies have indicated that duplication in the vasoactive intestinal peptide receptor-2 (VIPR2) gene confers the susceptibility to SCZ in Caucasians, but so far this finding has still not been confirmed in Chinese populations. In this study, we investigated the association between CNVs in VIPR2 and SCZ risk in an independent case–control study of Han Chinese using 1035 cases and 1535 controls. The CNVs were genotyped using the multiplex fluorescence competitive PCR method. In contrast with a common genotype (2-copy), a microduplication variant genotype (3-copy) was only carried by SCZ patients (4/1035). This finding indicated that CNVs in VIPR2 may impose a significantly increased risk of SCZ in Han Chinese (P=0.02646, OR=infinity, 95% CI=1.327–infinity). Thus, our results suggest that carriers of microduplication genotypes of VIPR2 are predisposed to SCZ in Han Chinese.

Genetic Susceptibility to Refractive Error: Association of Vasoactive Intestinal Peptide Receptor 2 (VIPR2) with High Myopia in Chinese

Myopia is the most common ocular disease worldwide. We investigated the association of high myopia with the common single nucleotide polymorphisms (SNPs) of five candidate genes – early growth response 1 (EGR1), v-fos FBJ murine osteosarcoma viral oncogene homolog (FOS), jun oncogene (JUN), vasoactive intestinal peptide (VIP), and vasoactive intestinal peptide receptor 2 (VIPR2). We recruited 1200 unrelated Chinese subjects with 600 cases (spherical equivalent ≤−8.00 diopters) and 600 controls (spherical equivalent within ±1.00 diopter). A discovery sample set was formed from 300 cases and 300 controls, and a replication sample set from the remaining samples. Tag SNPs were genotyped for the discovery sample set, and the most significant haplotypes and their constituent SNPs were followed up with the replication sample set. The allele and haplotype frequencies in cases and controls were compared by logistic regression adjusted for sex and age to give P a values, and multiple comparisons were corrected by permutation test to give P aemp values. Odd ratios (OR) were calculated accordingly. In the discovery phase, EGR1, JUN and VIP did not show any significant association while FOS and VIPR2 demonstrated significant haplotype association with high myopia. In the replication phase, the haplotype association for VIPR2 was successfully replicated, but not FOS. In analysis combining both sample sets, the most significant association signals of VIPR2 were the single marker rs2071625 (P a = 0.0008, P aemp = 0.0046 and OR = 0.75) and the 4-SNP haplotype window rs2071623-rs2071625-rs2730220-rs885863 (omnibus test, P a = 9.10e-10 and P aemp = 0.0001) with one protective haplotype (GGGG: P aemp = 0.0002 and OR = 0.52) and one high-risk haplotype (GAGA: P aemp = 0.0027 and OR = 4.68). This 4-SNP haplotype window was the most significant in all sample sets examined. This is the first study to suggest a role of VIPR2 in the genetic susceptibility to high myopia. EGR1, JUN, FOS and VIP are unlikely to be important in predisposing humans to high myopia.

Effects of VIP on Corneal Reconstitution and Homeostasis followingPseudomonas aeruginosaInduced Keratitis

Studies from our laboratory have demonstrated that vasoactive intestinal peptide (VIP) directly converts the normally susceptible C57BL/6J (B6) mouse to resistant after ocular infection through modulation of the inflammatory response. This study examines mechanisms by which VIP influences the healing phase following infection--specifically reconstitution of the extracellular matrix (ECM). B6 mice received daily intraperitoneal (IP) injections of VIP, while control mice were similarly injected with sterile phosphate buffered saline (PBS). Real-time RT-PCR, ELISA, and immunofluorescent staining were used to assess the effects of VIP treatment on ECM molecule expression after Pseudomonas aeruginosa-induced keratitis. We also compared the effect of VIP treatment on lipopolysaccharide (LPS)-stimulated B6- and BALB/c-derived fibroblasts. In vivo analyses revealed that VIP treatment of P. aeruginosa-infected B6 corneas led to a significant increase in ECM molecules associated with healing/homeostasis, while those associated with ECM degradation were significantly down-regulated when compared to wild-type (WT) controls. In vitro studies revealed that VIP treatment of lipopolysaccharide-stimulated fibroblasts derived from susceptible B6 and resistant BALB/c mice expressed distinct differences in ECM molecule expression, whereby the latter expressed higher levels of ECM molecules aimed at reconstitution. Furthermore, differential expression of VIP receptor-1/VIP receptor-2 (VIPR1/VIPR2) was observed between B6 and BALB/c after VIP treatment of LPS-stimulated fibroblasts. VIP treatment functions to enhance ECM reconstitution, which appears to be carried out in large part by fibroblasts via VIPR2. Overall, the data from this study suggest that VIP not only regulates disease pathogenesis, but also functions to restore integrity of the corneal stroma.