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Abstract Introduction: The endoparasite Dendrogaster argentinensis infects the intertidal brooder sea star Anasterias antarctica. This sea-star species is in the highest trophic level in the Beagle Channel. Objective: To study the effects of parasitism by D. argentinensis on the fitness and reproduction of A. antarctica. Methods: Adults from the brooder sea-star were collected from the rocky intertidal of Ensenada Zaratiegui bay (54°51' S & 68°29' W), Argentina. Eight seasonal samplings were performed (four seasons in two years) in the upper and low intertidal. During dissection, parasites were removed, and all organs were extracted and weighed separately. Results: Dendrogaster argentinensis prevalence was the highest for the region (20.4 %). Parasitized individuals were more frequent in the low intertidal in all seasons, with a higher difference in summer, where it is likely that the higher temperatures and strong winds could make the upper intertidal more challenging for a parasitized individual. Five parasitized individuals were castrated. Generally, the gonadal (GI) and somatic (pyloric caeca, PCI; stomach, SI; body wall, WI) indexes were lower in parasitized than non-parasitized individuals. Conclusions: Parasitism by D. argentinensis negatively affects A. antarctica condition. It affects reproduction because it reduces the GI, and can also produce castration. The parasite competes for the sea-stars' energetic resources, also decreasing the individual's capacity for feeding (reduced stomach) and growth (reduced body wall).
Resumen Introducción: El endoparásito Dendrogaster argentinensis infecta a la estrella de mar Anasterias antarctica, especie que se encuentra en el nivel trófico más alto del Canal Beagle. Objetivo: Estudiar los efectos del parasitismo de D. argentinensis en la condición fisiológica y reproducción de A. antarctica. Métodos: Adultos de la estrella de mar incubadora fueron recogidos del intermareal rocoso de la bahía Ensenada Zaratiegui (54°51' S & 68°29' W). Se realizaron ocho muestreos estacionales (cuatro temporadas en dos años) en el intermareal superior y bajo. Durante la disección, se removieron los parásitos, y todos los órganos, los cuales fueron pesados por separado. Resultados: La prevalencia de D. argentinensis fue la más alta de la región (20.4 %). Los individuos parasitados fueron más frecuentes en el intermareal bajo en todas las estaciones, siendo la mayor diferencia en verano, donde es probable que las temperaturas más altas y los fuertes vientos puedan hacer que el intermareal superior sea más desafiante para un individuo parasitado. Se observaron cinco individuos parasitados que estaban castrados. Generalmente, los índices gonadales (GI) y somáticos (ciego pilórico, estómago, y pared del cuerpo) fueron menores en los individuos parasitados que no parasitados. Conclusiones: El parasitismo de D. argentinensis afecta negativamente la condición fisiológica de A. antarctica. Afecta a la reproducción en términos de bajo GI y puede causar castración. El parásito compite por los recursos energéticos de las estrellas de mar, disminuyendo también la capacidad del individuo para alimentarse (reducción del estómago) y crecer (reducción de la pared del cuerpo).
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Animales , Parásitos/microbiología , Estrellas de Mar/parasitologíaRESUMEN
Cold and heat belong to the eight-principal syndrome differentiation of traditional Chinese medicine， which can reflect the rise and fall of Yin and Yang in the body and the Yin and Yang nature of the disease. At present， traditional Chinese medicine has an inconsistent understanding of cold and heat in acute coronary syndrome. The emphasis on pathogenic factors of cold and heat is biased， and the elements of cold and heat syndrome are not fully reflected in the scale. Therefore， the literature has been reviewed from the perspectives of etiology， pathogenesis， symptom elements， and test signs with drugs. From the perspective of etiology， both cold evil and heat evil can increase the risk of acute coronary syndrome. It was previously believed that acute coronary syndrome occurs frequently in cold climates such as winter and spring. Based on this understanding， hot weather can also induce acute coronary syndrome， and different temperatures have different effects on patients of different ages and with different underlying diseases. In addition， artificial pathogenic factors such as excessive consumption of cold food and refrigeration air conditioners were added. From the perspective of pathogenesis， on the basis of the traditional ''asthenia in origin and asthenia in superficiality'' and ''phlegm stagnation''， it is found that Yin-cold and fire-heat can both cause paralysis of the heart chakra and pain induced by the blockage. The pathogenesis of acute coronary syndrome characterized by heat stagnation and coldness featuring heartburn should be distinguished from gastroesophageal reflux disease. Moreover， the pathogenesis of Yin cold coagulation and pulse stagnation and wind obstruction are different. The acute coronary syndrome is in line with the wind characteristics of frequent changes and can be treated with wind medicine. From the perspective of syndrome elements， the syndrome elements such as cold condensation， heat accumulation， and toxicity are analyzed， and the use of basic syndrome elements and their combination forms facilitates clinical and scientific research. In addition， according to the test sign with the drug， it can be seen that the attributes of cold and heat of traditional Chinese medicine prescriptions for acute coronary syndrome can be explained according to the temperature-sensitive transient receptor potential （TRP） ion channel， thus proving the pathogenesis of cold and heat of acute coronary syndrome.
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Objective To investigate the therapeutic effects and mechanisms of Maxing Shigan Decoction on cough variant asthma(CVA)rats.Methods Sixty rats were randomly divided into normal group,model group,low and high dose groups of Maxing Shigan Decoction,and high-dose of Maxing Shigan Decoction + signal transducer and activator of transcription 3(STAT3)activator Colivelin(Col)group,12 rats in each group.Except for the normal group,the CVA model was constructed by intraperitoneal injection of ovalbumin combined with moxa fumigation in all other groups of rats.After the corresponding treatment,the rats were observed for signs and cough counts,airway resistance(RE)was detected by pulmonary function meter,eosinophils(EOS)were counted by Diff-Quik staining,histopathological features of the lungs and bronchial tubes were observed by hematoxylin-eosin(HE)staining method,and the lung tissues were detected by enzyme-linked immunosorbent assay(ELISA)for monocyte chemotactic protein 1(MCP-1),and tumor necrosis factor α(TNF-α),and the protein expression levels of interleukin 6(IL-6),STAT3,and transient receptor potential vanilloid-1 channel(TRPV1)were detected by Western Blot.Results Compared with the normal group,rats in the model group showed obvious asthma symptoms,severe inflammatory cell infiltration was seen in the lung tissue,bronchial epithelial cell necrosis,ciliated adhesion,mucus,and RE,EOS number,MCP-1 and TNF-α contents,and protein expression levels of IL-6,STAT3,TRPV1 were elevated(P＜0.05);compared with the model group,rats in the low-and high-dose groups of Maxing Shigan Decoction showed significant improvement in asthma symptoms,reduction in lung and bronchial injury,and dose-dependent reduction in RE,EOS number,MCP-1 and TNF-α contents,and protein expression levels of IL-6,STAT3,and TRPV1(P＜0.05);compared with the high-dose group of Maxing Shigan Decoction,the rats in the high-dose Maxing Shigan Decoction+Col group showed increased asthma,increased lung and bronchial injury,and increased RE,EOS number,MCP-1 and TNF-α contents,and protein expression levels of IL-6,STAT3,and TRPV1(P＜0.05).Conclusion Maxing Shigan Decoction can effectively improve cough variant asthma in rats,and its mechanism is related to the inhibition of IL-6/STAT3 signaling pathway and the high expression of TRPV1.
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Objective:Trigeminal neuralgia(TN)is a common neuropathic pain.Voltage-gated potassium channel(Kv)has been confirmed to be involved in the occurrence and development of TN,but the specific mechanism is still unclear.MicroRNA may be involved in neuropathic pain by regulating the expression of Kv channels and neuronal excitability in trigeminal ganglion(TG).This study aims to explore the relationship between Kv1.1 and miR-21-5p in TG with a TN model,evaluate whether miR-21-5p has a regulatory effect on Kv1.1,and to provide a new target and experimental basis for the treatment of TN. Methods:A total of 48 SD rats were randomly divided into 6 groups:1)a sham group(n= 12),the rats were only sutured at the surgical incision without nerve ligation;2)a sham+ agomir NC group(n=6),the sham rats were microinjected with agomir NC through stereotactic brain injection in the surgical side of TG;3)a sham+miR-21-5p agomir group(n=6),the sham rats were microinjected with miR-21-5p agomir via stereotactic brain injection in the surgical side of TG;4)a TN group(n=12),a TN rat model was constructed using the chronic constriction injury of the distal infraorbital nerve(dIoN-CCI)method with chromium intestinal thread;5)a TN+antagonist NC group(n=6),TN rats were microinjected with antagonist NC through stereotactic brain injection method in the surgical side of TG;6)a TN+miR-21-5p antagonist group(n=6),TN rats were microinjected with miR-21-5p antagonist through stereotactic brain injection in the surgical side of TG.The change of mechanical pain threshold in rats of each group after surgery was detected.The expressions of Kv1.1 and miR-21-5p in the operative TG of rats were detected by Western blotting and real-time reverse transcription polymerase chain reaction.Dual luciferase reporter genes were used to determine whether there was a target relationship between Kv1.1 and miR-21-5p and whether miR-21-5p directly affected the 3'-UTR terminal of KCNA1.The effect of brain stereotaxic injection was evaluated by immunofluorescence assay,and then the analogue of miR-21-5p(agomir)and agomir NC were injected into the TG of rats in the sham group by brain stereotaxic apparatus to overexpress miR-21-5p.The miR-21-5p inhibitor(antagomir)and antagomir NC were injected into TG of rats in the TN group to inhibit the expression of miR-21-5p.The behavioral changes of rats before and after administration were observed,and the expression changes of miR-21-5p and Kv1.1 in TG of rats after intervention were detected. Results:Compared with the baseline pain threshold,the facial mechanical pain threshold of rats in the TN group was significantly decreased from the 5th to 15th day after the surgery(P<0.05),and the facial mechanical pain threshold of rats in the sham group was stable at the normal level,which proved that the dIoN-CCI model was successfully constructed.Compared with the sham group,the expression of Kv1.1 mRNA and protein in TG of the TN group was down-regulated(both P<0.05),and the expression of miR-21-5p was up-regulated(P<0.05).The results of dual luciferase report showed that the luciferase activity of rno-miR-21-5p mimics and KCNA1 WT transfected with 6 nmol/L or 20 nmol/L were significantly decreased compared with those transfected with mimic NC and wild-type KCNA1 WT,respectively(P<0.001).Compared with low dose rno-miR-21-5p mimics(6 nmol/L)co-transfection group,the relative activity of luciferase in the high dose rno-miR-21-5p mimics(20 nmol/L)cotransfection group was significantly decreased(P<0.001).The results of immunofluorescence showed that drugs were accurately injected into TG through stereotaxic brain.After the expression of miR-21-5p in the TN group,the mechanical pain threshold and the expression of Kv1.1 mRNA and protein in TG were increased.After overexpression of miR-21-5p in the sham group,the mechanical pain threshold and the expression of Kv1.1 mRNA and protein in TG were decreased. Conclusion:Both Kv1.1 and miR-21-5p are involved in TN and miR-21-5p can regulate Kv1.1 expression by binding to the 3'-UTR of KCNA1.
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Objective To investigate the correlation between the expression of purinergic ligand-gated ion channel 7 receptor(P2X7R)and connective tissue growth factor(CTGF)in serum and cognitive function and clinical symptoms in patients with schizophrenia.Methods A total of 160 patients with schizophrenia who were diagnosed and treated in Department of Mental Intensive Care of Wuhan Wudong Hospital from January 2021 to January 2023 were collected as the observation group,and 160 healthy volunteers who underwent physical examinations were collected as the control group for the study.According to the Positive and Negative Symptom Scale(PANSS),patients were evaluated for their clinical and psychiatric symptoms(positive and negative symptoms,general pathological symptoms,and additional symptoms).The patients were grouped into a high score group(PANSS total score≥70 points,n=72)and a low score group(PANSS total score＜70 points,n=88).MATRICS consensus cognitive battery(MCCB)was applied to evaluate the cognitive abilities of patients;enzyme-linked immunosorbent assay(ELISA)was applied to detect serum P2X7R and CTGF levels.Spearman method was applied to analyze the correlation between serum P2X7R,CTGF levels and PANSS scores,and MCCB scores in patients with schizophrenia.Results Compared with the control group,the serum levels of P2X7R(610.71±107.83ng/L vs 384.78±80.62 ng/L)and CTGF(1.85±0.36μg/L vs 1.40±0.21μg/L)in the observation group were increased,with differences of statistical significance(t=21.226,13.658,all P＜0.05).The scores of variety items of MCBB of patients with schizophrenia in the observation group were lower than those in the control group,with differences of statistical significance(t=14.845～24.862,all P＜0.05),the positive symptom score(21.10±3.42score),negative symptom score(23.37±5.03 score),general pathological symptom score(39.48±8.11score),additional symptom score(8.26±1.22 score),and PANSS total score(92.21±12.50score)of schizophrenia patients in the high group were higher than those in the low group(13.65±3.04,15.62±3.91 score,30.14±6.15 score,5.20±0.94score,64.61±5.30score),with differences of statistical significance(t=14.576,10.964,8.280,17.915,18.764,all P＜0.05).The serum levels of P2X7R and CTGF in patients with schizophrenia in the high group were higher than those in the low group,with differences of statistical significance(t=12.233,5.923,all P＜0.05).The levels of serum P2X7R and CTGF in patients with schizophrenia were positively correlated with PANSS score(r=0.464～0.580,all P＜0.05),and negatively correlated with MCCB score[r=-0.603～-0.439,all P＜0.05].Conclusion The serum levels of P2X7R and CTGF in patients with schizophrenia are elevated,they are closely related to the clinical symptoms and cognitive function of patients.
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Objective:To investigate the protective mechanism of TRPV4 channel inhibitor on blood-brain barrier(BBB)damage after traumatic brain injury(TBI).Methods:The TBI rat model was established.TRPV4 channel inhibitor HC067047 or PKC-δ inhibitor Rottlerin was used to detect changes in BBB permeability,neurological function score,and the expression of microvascular endothelial tight junction proteins ZO-1 and ZO-2 in brain injury areas after TBI.Results:Compared with the Sham group,BBB permeability significantly increased,brain neurological function score significantly decreased,and the expression of ZO-1 and ZO-2 significantly decreased in TBI group(P＜0.05).Compared with the TBI group,after administration of HC067047 or Rottlerin,changes in BBB permeability,brain neurological function score,the expression of ZO-1 and ZO-2 were partially reversed(P＜0.05).Conclusions:TBI-induced BBB injury may be mediated by TRPV4 channel regulating PKC-δ signaling pathway to affect the expression of tight junction proteins ZO-1 and ZO-2.Inhibition of TRPV4 channel function or PKC-δ signal molecule can partially alleviate BBB damage induced by TBI.This study may provide new ideas for the treatment of clinical TBI.
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Objective To compare the safety and clinical efficacy of lesion removal combined with percutaneous pedicle screw fixation with classical posterior lesion removal in the treatment of lumbar brucelli spondylitis(LBS)by unilateral biportal endoscopic technique with transforaminal lumbar interbody fusion(UBE-LIF)technique.Methods The clinical data of 32 patients with LBS admitted by the Department of Spine and Orthopedics of Gansu Provincial Hospital of Traditional Chinese Medicine from January 2020 to January 2022 were retrospectively analyzed,and the clinical data of the 32 LBS patients were divided into 15 cases in the UBE-LIF group and 17 cases in the posterior group.The general data,surgery-related indexes,and postoperative pathological HE staining of the two groups were recorded and analyzed.The patients'clinical recovery was assessed according to their erythrocyte sedimentation rate(ESR)and C-reactive protein(CRP),low back pain visual analogue score(VAS),Japanese Orthopaedic Association(JOA)score,and Oswestry Dysfunction Index(ODI)preoperative,1 week after surgery,1,3,6 months and 1 year after surgery.Lumbar lordosis angle(LL)and intervertebral space height(DH)were measured by imaging before surgery and at the last follow-up,and intervertebral bone graft fusion was assessed using Suk grading criteria.Results Both groups successfully completed the operation and no serious postoperative complications occurred.There were no significant differences in gender,age,surgical segment,operation time,preoperative ESR and CRP,preoperative VAS,JOA score and ODI index,preoperative LL and DH(P＞0.05).The intraoperative blood loss,postoperative drainage,postoperative getting out of bed,and postoperative hospital stay in UBE-LIF group were significantly lower than those in the posterior group(P＜0.001).Pathological examination of diseased tissues was performed during surgery,all of which was consistent with brucellosis changes.Patients in both groups were followed up for 12-18 months,with an average of 14.8 months.The VAS,JOA score,and ODI index at all postoperative time points in the two groups were significantly improved compared with the preoperative period(P＜0.05).The difference between the two groups was significantly greater than that in the postoperative group:VAS score was lower in UBE-LIF group than in the posterior group(P＜0.01),CRP in both groups was higher than that in the preoperative group,and the elevation level was significantly lower in UBE-LIF group than in the posterior group(P＜0.001).There was no significant difference in ESR between the two groups compared with that before surgery(P＞0.05).There were no significant differences in VAS,JOA score,ODI index,CRP or ESR between the remaining time points after surgery(P＞0.05).At the last follow-up,imaging examination showed that the overall fusion rate of intervertebral bone graft in UBE-LIF group was 93.3％and 94.1％in the posterior group,without significant difference(x2=0.246,P=0.884).LL and DH were significantly improved in both groups compared with preoperative ones(P＜0.01),and the two groups did not significantly differ before and after surgery(P＞0.05).Conclusion Both surgical treatments for LBS are safe effect.Compared with posterior lesion removal bone graft fusion internal fixation,UBE-LIF technology combined with percutaneous pedicle screw internal fixation has the advantages of clear intraoperative vision,less blood loss,faster early postoperative recovery,and shorter postoperative hospital stay,and thus is a feasible surgical method for the minimally invasive treatment of LBS.
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BACKGROUND:Connexin 43(Cx43),which is thought to be engaged in the gap junction process and build the structural groundwork for the development of direct material signaling channels between cells,is crucial for maintaining the homeostatic balance of tissue metabolism.Recent research,however,has revealed fresh information about its distinct hemichannel function and highlighted the significance of its subcellular localization and self-fragmentation for cellular physiological activities and pathological processes. OBJECTIVE:To systematically summarize the molecular characteristics and expression of Cx43 in a variety of cells,concentrate on the pathological and physiological roles of channel-dependent Cx43 and channel-independent Cx43,and investigate the potential value in disease treatment by reviewing the pertinent literature in the database. METHODS:The Chinese and English keywords were"gap junction,connexin 43(Cx43),hemichannel,channel-dependent Cx43,channel-independent Cx43,extracellular vesicles(EVs),mitochondria,GJA1-20k",which were searched in PubMed and CNKI.Finally,81 articles were selected for review. RESULTS AND CONCLUSION:(1)The canonical role of Cx43 is to form a gap junction channel.Channel-dependent Cx43 has primarily involved in disease physiopathological processes by directly constituting gap junction channels,but full attention should be paid to the issue of its structural and functional integrity.Adhesion is a crucial characteristic of gap junctions,which are strongly associated with barrier-like diseases.(2)The non-canonical role of Cx43 is non-gap junction channel-dependent effect.In addition to being localized at the plasma membrane,inner mitochondrial membrane,extracellular vesicle surface,and other structures,Cx43 hexamer has also been found to play a role in positive pro-inflammatory mechanisms,mitochondrial functional metabolism,and targeted uptake of extracellular vesicles in inflammatory diseases.Selective shortened segments control mitochondrial homeostasis by encouraging the polymerization of peri-mitochondrial actin and are involved in the targeted translocation of full-length Cx43 to intracellular structural domains.(3)The development of targeted medicines and the solving of issues like the mechanism of seed cell transformation in tissue engineering-based therapies are both made possible by these two categories of impacts.The interactions of various types of Cx43,however,are frequently not fully taken into account in some of the existing original studies,which confuses the overall characteristics and skews the results.(4)It is necessary to systematically frame the physiological characteristics of Cx43 in different forms and its potential mechanisms in various diseases,so as to provide a reference for the exploration of the Cx43 integrity mechanism and the diagnosis and treatment of multiple diseases.
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BACKGROUND:Fluorosis is a disorder of enamel development caused by long-term intake of large amounts of fluoride during enamel development. OBJECTIVE:To further explore the molecular mechanism of dental fluorosis formation by screening the differentially expressed genes associated with calcium homeostasis in ameloblasts by transcriptome sequencing technology. METHODS:LS8 cells were treated with 0,0.4,0.8,1.6,3.2 and 6.4 mmol/L sodium fluoride(NaF)for 24,48 and 72 hours to observe the effects of different concentrations of NaF on the morphology,cell activity and intracellular Ca2+ concentration of LS8 cells.The differentially expressed genes were screened by transcriptome sequencing and validated. RESULTS AND CONCLUSION:After 24 hours of treatment,the cells treated with 0,0.4,and 0.8 mmol/L NaF were in good growth condition,with increased cell number and clear cell outline.When the NaF concentration was≥1.6 mmol/L,the cells were gradually shrunken and became smaller and the number of cells decreased with the increase of NaF concentration.After 48 and 72 hours of treatment,the number of cells increased in the 0,0.4 mmol/L NaF groups,while gradually decreased in the 0.8,1.6,3.2 mmol/L NaF groups,with rounded and smaller cell morphology.The cells in the 6.4 mmol/L NaF group were shrunken,rounded and suspended in the medium,with almost no adherent cells.When treated with the same concentration of NaF,LS8 cells were in optimal growth after 24 hours of treatment.Results from cell counting kit-8 assay showed that when treated with the same concentration of NaF,the cell activity decreased with the increase of treatment time;when the treatment time was the same,the cell activity decreased with the increase of NaF concentration.After 24 hours of treatment,the intracellular Ca2+ concentration increased with the increase of NaF concentration.Transcriptome sequencing analysis identified genes involved in the regulation of cellular calcium homeostasis:Hsp90b1,Canx,Calr,and Hspa5 that were significantly upregulated(P＜0.05)and Cacna1a that was significantly downregulated(P＜0.05).To conclude,the inhibitory effect of NaF on LS8 cell proliferation may be related to the abnormal increase in intracellular Ca2+ concentration,and the mechanism may be caused by the upregulation of the expression of protein processing and synthesis pathways Hsp90b1,Canx,Calr,and Hspa5 and the downregulation of the expression of calcium signaling pathway Cacna1a.
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BACKGROUND:Muscle weakness is a common symptom after coronavirus disease 2019(COVID-19)infection and affects the ability to perform daily activities in humans during recovery.Low-frequency pulsed magnetic field stimulation at a strength of 1.5 mT and a frequency of 3 300 Hz can enhance the maximal voluntary contraction and strength endurance of human skeletal muscle by inducing and activating classical transient receptor potential channel 1(TRPC1),which produces a series of pathological support effects on muscle tissue.It has not been studied whether this means will improve muscle weakness in patients recovering from COVID-19. OBJECTIVE:To select the low-frequency pulsed magnetic field for magnetic stimulation of lower limb muscle groups in patients with COVID-19,in order to observe the effect of this stimulation on the improvement of muscle weakness of lower limb muscle groups in patients with COVID-19 during the recovery period. METHODS:Fourteen patients infected with COVID-19(Omicron strain)positive for Innovita COVID-19 Ab Test(Colloidal Gold)and accompanied by muscle weakness were recruited and randomly divided into two groups:a test group receiving magnetic field stimulation and a control group receiving sham treatment,respectively.The total duration of the trial was 3 weeks.The test group was given low-frequency pulsed magnetic stimulation of the lower limbs every 48 hours and the control group was given the same intervention procedure as the test group but with sham stimulation.Patients in both groups were not informed whether the magnetic stimulation apparatus was running or not.Nine sessions were performed in both groups and the changes in the maximum voluntary contraction,explosive leg force and strength endurance of the local muscle groups of the lower limbs were subsequently observed in both groups. RESULTS AND CONCLUSION:Among the eight local muscle groups collected,seven local muscle groups in the test group showed an increase in the maximum voluntary contraction value after 3 weeks of low-frequency pulsed magnetic field stimulation.In the control group,there were only three muscle groups with improvement in the maximum voluntary contraction.The rate of improvement in the anterior and posterior muscle groups of the left leg in the test group was significantly higher than that in the control group.The longitudinal jump height and peak angular velocity of the knee joint in both groups were improved compared with the pre-test measurement,and the elevation rate of jumping height in the test group was higher than that in the control group.Under the fatigue condition,the decline rates of peak angular velocity of the knee joint and jumping height in the test group decreased significantly,while those in the control group did not change significantly.The above data confirmed that the low-frequency pulsed magnetic field stimulation with the intensity of 1.5 mT and frequency of 3 300 Hz could improve the muscle strength of more local muscle groups in the lower limbs of patients with COVID-19 during the recovery period compared with the human self-healing process,and the whole-body coordination ability and functional status based on explosive leg force of the legs could be significantly improved.Therefore,low-frequency pulsed magnetic field stimulation can be used as an effective,non-exercise rehabilitation tool to improve muscle weakness in the lower limbs of patients with COVID-19.
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Objective:To investigate the effects and mechanism of exosome (EXO)-loaded kringle V11 (KV11) delivery on corneal neovascularization (CNV).Methods:KV11 was bound to the surface of endothelial cell-derived exosomes by using CP05, an EXO-targeting anchoring peptide, to produce EXO-KV11.The binding efficiency and optimal concentration ratio were determined using the Apogee flow system.A total of 100 8-week-old healthy male SPF grade SD rats were selected, 10 of which were randomly selected as a normal control group without any treatment.The CNV model was established by alkali burn in the other 90 rats, which were randomly divided into three groups, EXO-KV11 group, KV11 group, and normal saline group by the random number table method, with 30 rats in each group.Each group was injected subconjunctivally with 100 μl of EXO-KV11 (25 μg), KV11 (25 μg), or normal saline every other day from the first day after the alkali burn, respectively.The CNV of rats was observed on days 1, 4, 7, and 14 after alkali burn.The CNV area was calculated by ventricular perfusion with fluorescein isothiocyanate-dextran (FITC-dextran) and corneal angiography.The amount of CNV lumen was observed by hematoxylin and eosin staining.The distribution of CD31 in rat corneas was determined by immunohistochemical method.The expression levels of voltage-dependent anion channel 1(VDAC1), endoplasmic reticulum stress, autophagy and apoptosis-associated proteins were detected by Western blot.This study was approved by the Animal Ethics Committee of Peking University People's Hospital (No.20210019). All animal procedures complied with the regulations of the Vision and Ophthalmology Association and the Animal Protection and Use Committee of Peking University.Results:The optimal concentration ratio of KV11 to EXO was 4∶1 and the binding affinity reached up to 87.5% by Apogee flow cytometers.On days 7 and 14 after alkali burn, there were significant differences in CNV area among the four groups ( F=4.613, 15.590; both at P<0.05). On day 7 after alkali burn, the CNV area was smaller in EXO-KV11 group than in KV11 and normal saline groups, with statistically significant differences (both at P<0.05). On day 14 after alkali burn, the CNV area was smaller in EXO-KV11 and KV11 groups than in normal saline group, and smaller in EXO-KV11 group than in KV11 group, showing statistically significant differences (all at P<0.05). The results of quantitative analysis of corneal fluorescence mounts showed that the relative CNV fluorescence area of the normal saline group, KV11 group and EXO-KV11 group were (8.3±1.7)%, (5.2±1.6)%and (3.4±0.7)%, respectively, showing a statistically significant overall comparison difference ( F=11.735, P<0.01). The relative CNV fluorescence area was larger in KV11 and normal saline groups than in EXO-KV11 group, and larger in normal saline group than in KV11 group, showing statistically significant differences (all at P<0.05). On day 14 after alkali burn, massive neovascular lumens were observed in the matrix of the normal saline group.The number of neovascular lumens in KV11 group was smaller than that in normal saline group.The corneal structure appeared normal in EXO-KV11 group, and neovascular lumens were rare.Numerous CD31-positive cells were observed in the corneal stroma of the normal saline group, which formed into lumen structures.The number of lumens surrounded by CD31-positive cells in the corneal stroma was smaller in KV11 group than in normal saline group, and smaller in EXO-KV11 group than in KV11 group.There were significant differences in the relative expression levels of VDAC1, protein kinase R-like endoplasmic reticulum kinase (PERK), p62, cleaved caspase 3 among the four groups ( F=35.960, 8.947, 17.791, 101.168; all at P<0.01). The relative expression levels of VDAC1, PERK, p62, cleaved caspase 3 were higher in EXO-KV11 group than in KV11 and normal saline groups, showing statistically significant differences (all at P<0.001). There was no significant difference in the relative expression of microtubule-associated proteins 1A/1B light chain 3B (LC3B)Ⅱ/LC3BⅠ protein among all four groups ( F=0.445, P=0.727). Conclusions:EXO-KV11 can inhibit CNV more remarkably than KV11.EXO-KV11 inhibits CNV by promoting the expression of VDAC1 and PERK and suppressing the autophagic flux.
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Objective:To investigate the regulatory effect of transient receptor potential cation channel subfamily C member 3 (TRPC3) on the retina in oxygen-induced retinopathy (OIR) mice and biological behavior of human retinal vascular endothelial cells (HREC).Methods:A total of 32 healthy SPF grade 7-day-old C57BL/6 mice were selected and randomly divided into a control group and an OIR group by the random number table method, with 16 mice in each group.The control group received no special treatment, and the OIR model was established in the OIR group.On postnatal day 17 (PN17), the success of the model establishment was verified by immunofluorescence staining of the retinal patch.The in vitro cultured HREC were divided into a normal control group, a transfection reagent group, and a si-TRPC3 group.The normal control group received no special treatment, while the transfection reagent group and the si-TRPC3 group were transfected with transfection reagent or transfection reagent + si-TRPC3.The relative expression of TRPC3 mRNA was detected by real-time quantitative fluorescence PCR.The relative expressions of TRPC3, transcription factor NF-E2 related factor (Nrf2), and superoxide dismutase (SOD) proteins were determined by Western blot.HREC were further divided into a normal control group, a vascular endothelial growth factor (VEGF) group, a si-TRPC3 group, and a Pyr3 (TRPC3 channel inhibitor) group, which were cultured in complete medium, medium containing 20 ng/ml VEGF recombinant protein, medium containing 20 ng/ml VEGF recombinant protein (si-TRPC3 transfection for 72 hours), and medium containing 20 ng/ml VEGF recombinant protein+ 1 μmol/L Pyr3 for 48 hours, respectively.The proliferation ability of HREC was detected using cell counting kit 8 (CCK-8). The horizontal and vertical migration ability of cells were detected by cell scratch assay and transwell assay, respectively.This study followed the 3R principles of animal welfare and was approved by the Ethics Committee of Hebei Eye Hospital (No.2023LW04). Results:Pathological neovascular clusters with strong fluorescent staining appeared in the retina of OIR mice on PN17.The relative expressions of TRPC3 mRNA and protein in the retina of OIR mice were 2.057±0.244 and 1.517±0.290, respectively, significantly higher than 0.983±0.033 and 0.874±0.052 of control group ( t=6.165, 3.094; both at P<0.05). The relative expression levels of TRPC3 mRNA and protein were significantly lower, and the relative expression levels of Nrf2 and SOD proteins were higher in the si-TRPC3 group than in the normal control and transfection reagent groups, and the differences were statistically significant (all at P<0.05). The CCK-8 experiment results showed that the cell absorbance value was higher in the VEGF group than in the normal control group, and lower in the si-TRPC3 and Pyr3 groups than in the VEGF group, with statistically significant differences (all at P<0.05). The results of the cell scratch experiment showed that the lateral migration rate of VEGF group cells was higher than that of normal control group, while the lateral migration rate of si-TRPC3 group and Pyr3 group cells was lower than that of VEGF group, and the differences were statistically significant (all at P<0.05). The transwell experiment results showed that the number of stained cells in the VEGF group was higher than that in the normal control group, and the number of stained cells in the si-TRPC3 group and Pyr3 group was lower than that in the VEGF group, with statistically significant differences (all at P<0.05). Conclusions:Hypoxia induces increased TRPC3 expression in OIR mouse retina, and downregulation of TRPC3 inhibits HREC proliferation and migration.The mechanism is related to the activation of the Nrf2-related oxidative stress pathway.
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During the occurrence and development of various heart diseases,continuous deterioration of myocardial fibrosis leads to remodeling and dysfunction of the cardiac structure.As a newly discovered mechanically sensitive ion channel,Piezo1 has opened up a new field of research on cellular mechanical transduction.Piezo1 combines a fine force transducer with Ca2+ influx and participates in the regulation of cellular mechanical transduction,thereby regulating cellular biological functions.Recent studies have shown that the biomechanical changes induced by myocardial injury regulate the expression of Piezo1 in cardiomyocytes and cause an imbalance in calcium homeostasis,which plays an important role in the positive feedback loop of myocardial fibrosis.This review summarizes the theoretical basis and related studies of Piezo1 in regulating cardiac fibrosis and suggests that the Piezo1 channel may become a new target for the treatment of cardiac fibrosis,thereby providing a new research horizon for the prevention and treatment of cardiac fibrosis.
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AIM:To investigate the mechanism of Piezo1 channel activation promoting the increase in intra-cellular Ca2+ concentration([Ca2+]i)of rat coronary artery smooth muscle cells(CASMCs).METHODS:The primary CASMCs of SD rats were cultured,and the expression and subcellular localization of Piezo1 in the cells were observed by immunofluorescence staining.The Piezo1 and stromal interaction molecule 1(STIM1)in CASMCs were knocked down by siRNA transfection,and the expression levels of the proteins were detected by Western blot.Utilizing laser confocal mi-croscopy,the change of[Ca2+]i in CASMCs was detected by Fluo-4 AM fluorescent probes.RESULTS:It was confirmed by immunofluorescence staining that the expression of Piezo1 existed in primary rat CASMCs.Immunofluorescence staining also showed that Piezo1 was co-located with sarco-/endoplasmic reticulum Ca2+-ATPase 2(SERCA2),mitochondrial outer membrane protein TOM20 and nuclear membrane protein lamin B1.Western blot results showed that the protein expres-sion levels of STIM1 and Piezo1 were significantly down-regulated by siRNA transfection(P<0.05).Compared with con-trol group,Yoda1,the agonist of Piezo1,could increase the extracellular Ca2+ influx of CASMCs(P<0.01).However,the Ca2+ influx mediated by Yoda1 was not affected by the inhibition of L-type calcium channels.Treatment with Yoda1 in-creased the intracellular Ca2+ release of CASMCs(P<0.01).However,inhibition of calcium channels on endoplasmic re-ticulum,ryanodine receptor and inositol 1,4,5-triphosphate receptor,did not affect intracellular Ca2+ release mediated by Yoda1.After the Ca2+ in endoplasmic reticulum was emptied using thapsigargin(TG),Yoda1 also mediated the Ca2+ re-lease of other organelles in CASMCs(P<0.01).After inhibition of L-type calcium channels,treatment with store-operated calcium channel(SOCC)inhibitor BTP2 or knockdown of STIM1 led to the decrease in extracellular Ca2+ influx of CASMCs mediated by Yoda1(P<0.01).Treatment with TG increased the release of Ca2+ from the endoplasmic reticulum of CASMCs after knockdown of Piezo1(P<0.05),but the extracellular Ca2+ influx mediated by TG was not affected.After inhibition of L-type calcium channels and SOCC,knockdown of Piezo1 led to the decreases in intracellular Ca2+ release and extracellular Ca2+ influx induced by Yoda1(P<0.01).CONCLUSION:The Piezo1 agonist orchestrates the influx of extracellular Ca2+ by activating Piezo1 channels on the cell membrane and inducing the indirect activation of SOCC.More-over,it facilitates the release of Ca2+ from organelles.Consequently,these pathways synergistically elevate the[Ca2+]i of rat CASMCs.
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AIM:To investigate the effect of Bushen formulae(BHF)on bone metabolism and its possible mechanism in ovariectomized rats with high salt intake.METHODS:According to the random number table method,80 SPF-grade Sprague-Dawley rats were divided into sham group,ovariectomy(OVX)group,medium-high-salt diet(MSD)group,high-salt diet(HSD)group,BHF group,BHF with normal saline(BHF+NS)group,BHF+MSD group,and BHF+ HSD group,with 10 rats in each group.After modeling,different diets and BHF formula interventions were administered,and the concentrations of sodium chloride added to MSD group and HSD group were 2%(w/w)and 8%(w/w),respective-ly.The dose of BHF was 7.8 g·kg-1·d-1,once a day,and the treatment lasted for 12 weeks.Bone density,bone microar-chitecture,bone parameters,bone metabolism biomarkers,bone histopathological changes,the expression of epithelial sodium channel α(ENaCα),Na-Cl cotransporter(NCC),and voltage-gated chloride channel 3(ClC-3)proteins in bone tissue were detected in each group.RESULTS:Compared with sham group,the rats in OVX group had reduced bone density and destroyed bone microstructure.Compared with OVX group,the bone microstructure in MSD and HSD groups was more significantly damaged,while the levels of bone formation markers,bone glycoprotein(BGP)and type Ⅰ procolla-gen N-terminal peptide(PINP),were significantly increased in HSD group(P<0.05).Moreover,the levels of bone re-sorption markers,such as amino-terminal cross-linked telopeptides of type Ⅰ collagen(NTX),carboxy-terminal cross-linked telopeptides of type Ⅰ collagen(CTX)and tartrate-resistant acid phosphatase(TRACP),were significantly in-creased(P<0.05),indicating that bone metabolism was in high-conversion state.High-salt diet accelerated the structural destruction of bone trabeculae,and Western blot results showed that high-salt diet caused decreases in the protein expres-sion levels of ENaCα and ClC-3 and an increase in the protein expression level of NCC in femoral tissues(P<0.05).After BHF intervention,the expression of relevant ion channels caused by high salt could be regulated to different degrees.CONCLUSION:Bushen formulae could differentially regulate the expression of relevant ion channels ENaCα,ClC-3,and NCC induced by high salt to different degrees,which has certain ameliorative and therapeutic effects on the imbalance of bone metabolism.
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AIM:To examine the effects of borneol on inflammation and myocardial remodeling after myocar-dial infarction(MI)in mice,and to investigate the underlying mechanisms.METHODS:Eight-week-old wild-type(WT)C57BL/6 mice and transient receptor potential cation channel subfamily M member 8(TRPM8)gene knockout(TRPM8-/-)mice were randomly divided into sham and MI groups,and were subsequently treated with normal saline(control group)or borneol(borneol group)via gavage.Survival curves were plotted for WT and TRPM8-/-mice with MI treated with or with-out borneol.After 28 d,cardiac function of the mice was assessed through echocardiography,and haemodynamic indexes were evaluated using a multi-channel physiological instrument.Infarct size,myocardial hypertrophy and interstitial fibro-sis were assessed via pathological staining.In addition,inflammatory response in the peri-infarct region was detected.RE-SULTS:The TRPM8 expression was up-regulated in the peri-infarct region of the mice with MI(P＜0.05),and borneol had no effect on TRPM8 expression(P＞0.05).Borneol increased the survival rate,reduced the infarct size,inhibited car-diac remodeling and improved cardiac function in WT mice with MI(P＜0.05 or P＜0.01).However,it did not affect the survival rate,infarct size,myocardial hypertrophy,myocardial fibrosis or cardiac function in TRPM8-/-mice(P＞0.05).Furthermore,borneol reduced inflammatory cell infiltration and the expression of inflammatory cytokines,tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),IL-6 and monocyte chemotactic protein-1(MCP-1),in WT mice(P＜0.05 or P＜0.01)but not in TRPM8-/-mice(P＞0.05).CONCLUSION:Borneol attenuates inflammation,inhibits cardiac re-modeling and improves cardiac function in mice with MI via TRPM8.
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Objective:To evaluate the role of ryanodine receptor 2 (RyR2) in postoperative cognitive dysfunction (POCD) in aged rats.Methods:Sixty SPF healthy male Sprague-Dawley rats, aged 20 months, weighing 600-650 g, were divided into 3 groups ( n=20 each) using a random number table method: control group (group C), POCD group (group P) and dantrolene group (group D). A rat POCD model was prepared by closed reduction and internal fixation of left tibial fractures with sevoflurane anesthesia in P and D groups. RyR inhibitor dantrolene 2 mg/kg was injected via a tail vein at 30 min before surgery in group D. Morris water maze tests were conducted on day 1 before surgery and day 7 after surgery to evaluate the cognitive function. An open field test was conducted to detect the spontaneous motor function starting from day 7 after surgery. The rats were sacrificed after the end of Morris water maze tests and hippocampal tissues were taken for determination of the expression of RyR2 and cleaved caspase-3 (by Western blot), apoptosis rate and cytoplasmic calcium ion concentrations (by flow cytometry) and for microscopic examination of the pathological changes in hippocampal CA1 area (using HE staining). Results:There was no significant difference in the speed, distance and duration of stay in the center in the open field test among the three groups ( P>0.05). Compared with group C, the escape latency was significantly prolonged after surgery, the number of crossing the original platform was reduced, the expression of RyR2 and cleaved caspase-3 was up-regulated, and the neuronal apoptosis rate and cytoplasmic calcium ion concentration were increased ( P<0.05), and the pathological changes were found in the hippocampal CA1 area in group P. Compared with group P, the escape latency was significantly shortened after surgery, the number of crossing the original platform was increased, the expression of RyR2 and cleaved caspase-3 was down-regulated, and the neuronal apoptosis rate and cytoplasmic calcium ion concentration were decreased ( P<0.05), and the pathological changes were significantly reduced in the hippocampal CA1 area in group D. Conclusions:RyR2 activation is involved in the process of POCD in aged rats, which may be associated with increased calcium overload-induced hippocampal neuronal apoptosis.
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Objective:To evaluate the effect of gastrogin on AMP-activated protein kinase (AMPK)/transient receptor potential anchor protein 1 (TRPA1) signaling pathway in rats with neuropathic pain.Methods:Thirty-six SPF-grade healthy male Sprague-Dawley rats, aged 6-8 weeks, weighing 200-230 g, were divided into 3 groups ( n=12 each) using a random number table method: sham operation+ normal saline group (SHAM group), neuropathic pain+ normal saline group (NP group), and neuropathic pain+ gastrogin group (GAS group). Neuropathic pain was induced by chronic constrictive injury to sciatic nerve under 2% isoflurane anaesthesia. The sciatic nerve was only exposed but not ligated in SHAM group. Gastrogin 100 mg/kg was intraperitoneally injected for 14 consecutive days after developing the model in GAS group, while the equal volume of normal saline was given instead in SHAM and NP groups. The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured at 1 day before developing the model (T 0) and 1, 3, 5, 7, 10 and 14 days after developing the model (T 1-6). The rats were anesthetized and sacrificed following the measurement of pain thresholds at T 4 and T 6. The lumbar segment (L 4-6) of the spinal cord was removed for determination of TRPA1 mRNA expression (by quantitative real-time polymerase chain reaction), expression of TRPA1, AMPK and p-AMPK (by Western blot), expression of TRPA1 (by immunofluorescence staining) and expression of tumor necrosis-alpha(TNF-α), interleukin-1beta(IL-1β) and c-fos (by immunohistochemistry). Results:Compared with SHAM group, MWT and TWL were significantly decreased at T 1-6, the expression of TRPA1 mRNA, TRPA1, TNF-α, IL-1β and c-fos was up-regulated, the expression of p-AMPK was down-regulated ( P<0.05), and no significant change was found in AMPK expression in NP group ( P>0.05). Compared with NP group, MWT at T 3-6 and TWL at T 2-6 were significantly increased, the expression of TRPA1 mRNA, TRPA1, TNF-α, IL-1β and c-fos was down-regulated, and p-AMPK expression was up-regulated ( P<0.05), and no significant change was found in AMPK expression in GAS group ( P>0.05). Conclusions:The mechanism by which gastrogin reduces neuropathic pain may be related to modulating the expression of the AMPK/TRPA1 signaling pathway in rats.
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External physicochemical factors and emotional changes often lead to intermittent facial flushing in patients with rosacea, accompanied by burning, stinging, and itching sensations. Many studies have demonstrated that neurovascular dysfunction and neurogenic inflammation induced by neuropeptides released following the activation of ion channels were associated with the occurrence of rosacea. This review summarizes research progress on the role of ion channels in the pathogenesis of rosacea and provides evidence for further research on ion channels as potential therapeutic targets for rosacea.
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Fascia,the initial response site for mechanical stimulation in manipulations,is also the target of the effect of manipulations.As the essence of manipulation is"force",how mechanical stimuli are transduced into neuroelectric and biochemical signals in the fascia and how physical and chemical signals of the fascia initiate the mechanical stimulation effect are the common key questions in the study of the principle of manipulation.The physical changes in the fascial connective tissue caused by the manipulation,such as the deformation and displacement of the fascial tissue,can act on the nerve end receptors in the fascial layer and generate neural electrical signals;they can also activate the mechanoreceptors on the fascial cell membrane and convert mechanical signals into chemical signals via mechanosensitive ion channel transduction,triggering a physicochemical coupling response in the fascial microenvironment and producing mechanical stimulation through neuro-endocrine-immune system pathways.The"mechanical force of manipulation"in the fascia is transmitted through the meridian to facilitate the body's perception and transmission of mechanical stimulation signals,indicating that the fascia is the"sensor"of coupled response to the physicochemical information of mechanical stimulation of manipulation.