Adenyl cyclase activator forskolin protects against Huntington's disease-like neurodegenerative disorders

Long term suppression of succinate dehydrogenase by selective inhibitor 3-nitropropionic acid has been used in rodents to model Huntington＇s disease where mitochondrial dysfunction and oxidative damages are primary pathological hallmarks for neuronal damage. Improvements in learning and memory abilities, recovery of energy levels, and reduction of excitotoxicity damage can be achieved through activation of Adenyl cyclase enzyme by a specific phytochemical forskolin. In this study, intraperitoneal administration of 10 mg/kg 3-nitropropionic acid for 15 days in rats notably reduced body weight, worsened motor cocordination（grip strength, beam crossing task, locomotor activity）, resulted in learning and memory deficits, greatly increased acetylcholinesterase, lactate dehydrogenase, nitrite, and malondialdehyde levels, obviously decreased adenosine triphosphate, succinate dehydrogenase, superoxide dismutase, catalase, and reduced glutathione levels in the striatum, cortex and hippocampus. Intragastric administration of forskolin at 10, 20, 30 mg/kg dose-dependently reversed these behavioral, biochemical and pathological changes caused by 3-nitropropionic acid. These results suggest that forskolin exhibits neuroprotective effects on 3-nitropropionic acid-induced Huntington＇s disease-like neurodegeneration.

Adenylate cyclase activates the cAMP signalling pathway to enhance platelet-rich plasma-treated Achilles tendon disease,a theoretical bioinformatics-based study

The effectiveness of platelet-rich plasma(PRP)for the treatment of Achilles tendon disorders still needs to be evaluated through a series of prospective studies,but genomic analysis can reveal the existence of complementary PRP treatment options.Based on the 96 platelet activation-related genes in the Kyoto Encyclopedia of Genes and Genomes(KEGG)database,we performed Gene Ontology functional enrichment analysis and KEGG enrichment analysis,pathway correlation analysis,and enrichment mapping to determine the enrichment results of the gene set enrichment analysis and found that the cAMP signalling pathway may be the key to enhancing the effectiveness of PRP treatment.The cAMP signalling pathway interacts with the Rap1 signalling pathway and cGMPPKG signalling pathway to mediate the entire pathophy-siological process of Achilles tendon disease.Moreover,ADCY1-9 may be the key to the activation of the cAMP signalling network.Further based on the data in the Gene Expression Omnibus database,it was found that ADCY4 and ADCY7 may be the players that play a major role,associated with the STAT4-ADCY4-LAMA5 axis and the GRbeta-ADCY7-SEMA3C axis,which is expected to be a complementary target for enhancing the efficacy of PRP in the treatment of Achilles tendon disease.

Characterization of two halophilic adenylate cyclases from Thermobifida halotolerans and Haloactinopolyspora alba

Adenylate cyclase(AC)is the key enzyme that catalyzes the formation of cAMP from ATP.In this study,we discovered two novel class Ⅲ ACs with a halophilic property from Thermobifida halotolerans DSM 44931(ThAC)and Haloactinopolyspora alba DSM 45211(HaAC),respectively.The recombinant ThAC and HaAC were expressed in Escherichia coli with molecular weights of 36.1 and 36.0 kDa respectively.The presence of 2500 and 2200 mmol
L^(-1)1 NaCl significantly enhanced the enzyme activities of ThAC and HaAC,with 22-fold and 7.4-fold higher activities compared to those without NaCl,respectively.Several divalent metal ions were found to activate the recombinant ACs to different extents,and the optimal metal ion was Mg^(2+)for both ThAC and HaAC with concentrations of 80 mmol·L^(-1) and 40 mmol·L^(-1) respectively.Purified ThAC and HaAC had the optimal specific activities((4.59±0.35)×10^(4) and(7.76±0.52)×10^(4) U·mg^(-1))and catalytic efficiency(4.47 and 5.30 L·mmol^(-1)·s^(-1))at 45
℃ and 40
℃ respectively,while the optimum pH of both two recombinant ACs was 10.0.This is the first report of the halophilic Class III ACs,which could make new contributions to explore and study ACs for further associated investigations.

Effects of Yulangsan polysaccharide on monoamine neurotransmitters, adenylate cyclase activity and brain-derived neurotrophic factor expression in a mouse model of depression induced by unpredictable chronic mild stress

The present study established a mouse model of depression induced by unpredictable chronic mild stress. The model mice were treated with Yulangsan polysaccharide （YLSPS; 150, 300 and 600 mg/kg） for 21 days, and compared with fluoxetine-treated and normal control groups. Enzyme-linked immunosorbent assay, radioimmunity and immunohistochemical staining showed that following treatment with YLSPS （300 and 600 mg/kg）, monoamine neurotransmitter levels, prefrontal cortex adenylate cyclase activity and hippocampal brain-derived neurotrophic factor expression were significantly elevated, and depression-like behaviors were improved. Open-field and novelty-suppressed feeding tests showed that mouse activity levels were increased and feeding latency was shortened following treatment. Our results indicate that YLSPS inhibits depression by upregulating monoamine neurotransmitters, prefrontal cortex adenylate cyclase activity and hippocampal brain-derived neurotrophic factor expression.

Efficient production of cyclic adenosine monophosphate from adenosine triphosphate by the N-terminal half of adenylate cyclase from Escherichia coli

In this study,we aimed at developing an efficient biocatalytic process for bio-production of cyclic adenosine monophosphate(c AMP)from adenosine triphosphate(ATP).First,adenylate cyclase from Escherichia coli MG1655(EAC)and Bordetella Pertussis(BAC)were expressed in E.coli BL21(DE3)and comparatively analyzed for their activities.As a result,EAC from E.coli MG1655 exhibited a higher activity.However,amount of EAC were obtained in an insoluble form.Therefore,we expressed the first 446 amino acids of EAC(EAC446)to avoid the inclusion body.The effects of induction temperature,incubation time,and incubation p H were further evaluated to improve the expression of EAC446.Subsequently,the reaction process for the production of c AMP with ATP as a starting material was investigated.As none of c AMP was detected in the whole-cell based biocatalytic process,the reaction catalyzed by the crude enzyme was determined for c AMP production.What's more,the reaction temperature,reaction p H,metal ion additives and substrate concentration was optimized,and the maximum c AMP production of 18.45 g·L^-1was achieved with a yield of 95.4%after bioconversion of 6 h.

Effects of pituitary adenylate cyclase activating polypeptide on CD4^+/CD8^+T cell levels after traumatic brain injury in a rat model

BACKGROUND:The effect of pituitary adenylate cyclase activating polypeptide(PACAP)during traumatic brain injury(TBI) and whether it can modulate secondary injury has not been reported previously.The present study evaluated the potential protective effects of ventricular infusion of PACAP in a rat model of TBI.METHODS:Male Sprague Dawley rats were randomly divided into 3 treatment groups(n=6,each):sham-operated,vehicle(normal saline)^+TBI,and PACAP^+TBI.Normal saline or PACAP(1 ug/5uL) was administered intracerebroventricularly 20 minutes before TBI.Right parietal cortical contusion was produced via a weight-dropping method.Brains were extracted 24 hours after trauma.Histological changes in brains were examined by HE staining.The numbers of CD4^+ and CD8^+ T cells in blood and the spleen were detected via flow cytometry.RESULTS:In injured brain regions,edema,hemorrhage,inflammatory cell infiltration,and swollen and degenerated neurons were observed under a light microscope,and the neurons were disorderly arrayed in the hippocampi.Compared to the sham group,average CD4^+ CD8" lymphocyte counts in blood and the spleen were significantly decreased in rats that received TBI^+vehicle,and CD4^+ CD8^+ were increased.In rats administered PACAP prior to TBI,damage was attenuated as evidenced by significantly increased CD4^+,and decreased CD8^+,T lymphocytes in blood and the spleen.CONCLUSION:Pretreatment with PACAP may protect against TBI by influencing periphery T cellular immune function.

No association between a polymorphism of the adenylate cyclase type IX gene and major depressive disorder in the Chinese Han population

Previous studies have demonstrated that a missense single-nucleotide polymorphism variant （2316A〉G;rs2230739）of the adenylate cyclase type IX gene was associated with bipolar disorder and affective disorder.We determined genotype and allele frequencies using a ligase detection reaction method in 315 patients with major depressive disorder and 278 unrelated, sex-matched healthy control subjects.We did not detect any statistically significant differences in genotype and allele frequencies between patients and healthy control subjects.Furthermore,we found no significant difference between genders in major depressive disorder,nor between patients and controls in the same gender.These results suggest that 2316A〉G（rs2230739）may not be a risk factor for increasing susceptibility to major depressive disorder in the Chinese Han population.

Low-power laser irradiation inhibits arecoline-induced fibrosis:an in vitro study

Oral submucous fibrosis （OSF） is a potentially malignant disorder that is characterized by a progressive fibrosis in the oral submucosa. Arecoline, an alkaloid compound of the areca nut, is reported to be a major aetiological factor in the development of OSF. Low-power laser irradiation （LPLI） has been reported to be beneficial in fibrosis prevention in different damaged organs. The aim of this study was to investigate the potential therapeutic effects of LPLI on arecoline-induced fibrosis. Arecoline- stimulated human gingival fibroblasts （HGFs） were treated with or without LPLI. The expression levels of the fibrotic marker genes alpha-smooth muscle actin （a-SMA） and connective tissue growth factor （CTGF/CCN2） were analysed by quantitative real- time reverse transcription polymerase chain reaction （RT-PCR） and western blots. In addition, the transcriptional activity of CCN2 was further determined by a reporter assay. The results indicated that arecoline increased the messenger RNA and protein expression of CCN2 and a-SMA in HGF. Interestingly, both LPLI and forskolin, an adenylyl cyclase activator, reduced the expression of arecoline-mediated fibrotic marker genes and inhibited the transcriptional activity of CCN2. Moreover, pretreatment with SQ22536, an adenylyl cyclase inhibitor, blocked LPLI＇s inhibition of the expression of arecoline-mediated fibrotic marker genes. Our data suggest that LPLI may inhibit the expression of arecoline-mediated fibrotic marker genes via the cAMP signalling pathway.

Effects of Activated ACM on Expression of Signal Transducers in Cerebral Cortical Neurons of Rats

To explore the roles of astrocytes in the epileptogenesis, astrocytes and neurons were isolated, purified and cultured in vitro from cerebral cortex of rats. The astrocytes were activated by ciliary neurotrophic factor (CNTF) and astrocytic conditioned medium (ACM) was collected to treat neurons for 4, 8 and 12 h. By using Western blot, the expression of calmodulin dependent protein kinase Ⅱ (CaMKⅡ), inducible nitric oxide synthase (iNOS) and adenylate cyclase (AC) was de- tected in neurons. The results showed that the expression of CaMKⅡ, iNOS and AC was increased significantly in the neurons treated with ACM from 4 h to 12 h (P<0.05), and that of iNOS and AC peaked at 8 h and 12 h respectively. It was suggested that there might be some epileptogenic factors in the ACM and such signal pathways as NOS-NO-cGMP, Ca2+?CaM-CaMKⅡ and AC-cAMP-PKA might take part in the signal transduction of epileptogenesis.

Changes of Adenylate Cyclase and Guanylate Cyclase in the Frontal Cortex,Lenticula,Corpus Amygdaloideum,and Hippocampus in Morphine‑dependent Rats

To detect the changes of adenylate cyclase(AC)and guanylate cyclase(GC)in the four cerebral regions that are concerned with psychogenic dependence of morphine in rats,including the frontal cortex,lenticula,corpus amygdaloideum,and hippocampus.To discuss the relation between the expressions of AC and GC with the psychogenic dependence on morphine.Different periods of morphine‑dependent rat models were established,and enzyme histochemistry was used to detect the variations of AC and GC in four cerebral regions.Compared with the control group,AC and GC in all the morphine‑dependent groups increased.The data indicated that the amounts of AC and GC were significantly different between the morphine‑dependent groups and the control group when tested at periods of 1 week,2 weeks,4 weeks,and 8 weeks(P<0.05 or P<0.01).There were significant differences when comparing the 1‑week group with the 2‑week,4‑week,and 8‑week groups(P<0.05 or P<0.01).There were significant differences when comparing the 2‑week dependent group with the 4‑week dependent group or the 8‑week dependent group(P<0.05 or P<0.01).The activities of AC and GC increased in four cerebral regions of morphine‑dependent rats.The psychogenic dependence on morphine appears to be closely linked to the upgrade of AC and GC.

基于PACAP-cAMP信号通路研究电针治疗骶上脊髓损伤后逼尿肌反射亢进型膀胱的效应机制

Objective:To elucidate the underlying mechanism and effect of electroacupuncture(EA)on the neurogenic bladder following suprasacral spinal cord injury(ssCI).A rat model of detrusor hyperreflexia after SsCI was established to examine the urodynamics,detrusor muscle tissue morphology,the protein and mRNA expression levels of pituitary adenylate cyclase activating peptide(PACAP)and its receptor PAC1R,and cyclic adenosine monophosphate(cAMP)content in the detrusor muscle with a focus on the PACAPcAMP signaling pathway.Method:A total of 72 female SD rats were randomized into control group and sham operation group(n=12 per group)by using a random number table.The remaining 48 rats were established into the model of detrusor hyperreflexia after SsCI.After successful modeling,these rats were randomly assigned to model,EA,and EA+PACAP6-38 groups(n=12 per group).The unsuccessful modeled rats were used for exploratory observation.For the rats in EA group,"Ciliao(BL32)""Zhongji(CV3)",and"Sanyinjiao(SP6)"were needled and stimulated by EA.The PACAP receptor antagonist PACAP6-38 was administered intraperitoneally in the EA+PACAP6-38 group before EA,and EA was applied for seven consecutive days.After treatment,the urodynamics of the rats were analyzed,and hematoxylin and eosin staining was used to examine rat bladder detrusor tissue morphology.The expressions of PACAP-38 and PAC1R were detected by immunohistochemistry and Western blot.The mRNA expression levels of PACAP-38 and PAC1R were examined by RT-qPCR,while cAMP content was detected by ELISA.Results:(1)Compared with sham operation group,it was exhibited disarray in the transitional epithelium cells of the bladder in the modeled rats.The intercellular space was significantly widened,accompanied by inflammatory cell infiltration and noticeable tissue edema.Both the bladder initial pressure and leak point pressure of the rats were higher(P<0.01),whereas the maximum cystometric capacity and bladder compliance were lower(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,together with the cAMP content,were lower(P<0.05).(2)Compared with the model rats,the EA group showed reduced inflammatory response in the detrusor muscle tissue,with decreased monocyte infiltration and less severe tissue edema.The bladder smooth muscle cells exhibited increased integrity,and there was decreased cellular tissue edema,inflammatory cell infiltration,and fibroplasia.The bladder initial pressure and leak point pressure were lower(P<0.05),while the maximum cystometric capacity and bladder compliance were higher(P<0.01).The protein and mRNA expression levels of PACAP-38 and PAC1R in the detrusor muscle,along with the cAMP content,were higher(P<0.05).(3)Compared to the EA group,the EA+PACAP6-38 group showed a less organized arrangement of muscle fibers in the detrusor muscle tissue,larger intercellular space,monocyte infltration,and considerable tissue edema.The changes in bladder initial pressure and leak point pressure were not significant(P>0.05),while the maximum cystometric capacity and bladder compliance were lower(P<0.05).The changes in the protein and mRNA expressions of PACAP-38 within the detrusor muscle were not signifcant(P>0.05),whereas the protein and mRNA expressions of PAC1R were reduced(P<0.05),and the cAMP content within the detrusor muscle was lower(P<0.05).Conclusion:EA can ameliorate the uninhibited contractile condition of the detrusor muscle in the bladder following SSCI.By mediating the PACAP-cAMP signaling pathway,it reduces the pathological damage to the detrusor muscle,thereby improving bladder function.

Neuroprotective Effects of Brain-Gut Peptides: A Potential Therapy for Parkinson’s Disease

Parkinson's disease(PD) is the second most common neurodegenerative disease and is typically associated with progressive motor and non-motor dysfunctions.Currently, dopamine replacement therapy is mainly used to relieve the motor symptoms, while its long-term application can lead to various complications and does not cure the disease. Numerous studies have demonstrated that many brain-gut peptides have neuroprotective effects in vivo and in vitro, and may be a promising treatment for PD. In recent years, some progress has been made in studies on the neuroprotective effects of some newly-discovered braingut peptides, such as glucagon-like peptide 1, pituitary adenylate cyclase activating polypeptide, nesfatin-1, and ghrelin. However, there is still no systematic review on the neuroprotective effects common to these peptides. Thus,here we review the neuroprotective effects and the associated mechanisms of these four peptides, as well as other brain-gut peptides related to PD, in the hope of providing new ideas for the treatment of PD and related clinical research.