ADT-OH inhibits malignant melanoma metastasis in mice via suppressing CSE/CBS and FAK/Paxillin signaling pathway.

Hydrogen sulfide (H2S) is widely recognized as the third endogenous gas signaling molecule and may play a key role in cancer biological processes. ADT-OH (5-(4-hydroxyphenyl)-3H-1,2-dithiocyclopentene-3-thione) is one of the most widely used organic donors for the slow release of H2S and considered to be a potential anticancer compound. In this study, we investigated the antimetastatic effects of ADT-OH in highly metastatic melanoma cells. A tail-vein-metastasis model was established by injecting B16F10 and A375 cells into the tail veins of mice, whereas a mouse footpad-injection model was established by injecting B16F10 cells into mouse footpads. We showed that administration of ADT-OH significantly inhibited the migration and invasion of melanoma cells in the three different animal models. We further showed that ADT-OH dose-dependently inhibited the migration and invasion of B16F10, B16F1 and A375 melanoma cells as evaluated by wound healing and Transwell assays in vitro. LC-MS/MS and bioinformatics analyses revealed that ADT-OH treatment inhibited the EMT process in B16F10 and A375 cells by reducing the expression of FAK and the downstream response protein Paxillin. Overexpression of FAK reversed the inhibitory effects of ADT-OH on melanoma cell migration. Moreover, after ADT-OH treatment, melanoma cells showed abnormal expression of the H2S-producing enzymes CSE/CBS and the AKT signaling pathways. In addition, ADT-OH significantly suppressed the proliferation of melanoma cells. Collectively, these results demonstrate that ADT-OH inhibits the EMT process in melanoma cells by suppressing the CSE/CBS and FAK signaling pathways, thereby exerting its antimetastatic activity. ADT-OH may be used as an antimetastatic agent in the future.

Screening for upper gastrointestinal cancers with magnetically controlled capsule gastroscopy: a feasibility study.

BACKGROUND: The medical consortium is an intensive and disease-specific association that integrates tertiary public hospitals and medical examination centers in China. We aimed to evaluate the feasibility of the medical consortium for screening upper gastrointestinal (GI) cancers (MCSC) by magnetically controlled capsule gastroscopy (MCCG). METHODS: 6627 asymptomatic subjects underwent MCCG as part of health check-ups in the MCSC between March and November 2018. Relevant clinical data were collected and analyzed. RESULTS: The MCSC detected 32 patients with upper GI cancer (0.48 %) confirmed by pathology. The detection rate of early gastric cancer was 16.67 % (4 /24). Gastric polyps, ulcers, and submucosal tumors were found in 15.54 %, 3.76 %, and 3.17 % of subjects, respectively. The whole GI preparation and operation process were well tolerated. CONCLUSIONS: The MCSC was a feasible model for upper GI cancer screening, especially for asymptomatic subjects. Further prospective studies with better operational quality control are warranted.

Expression of microRNA-328 Functions as a Biomarker for Recurrence of Early Gastric Cancer (EGC) After Endoscopic Submucosal Dissection (ESD) by Modulating CD44.

BACKGROUND This study investigated the molecular mechanism of the effect of CD44 on the recurrence of EGC after ESD, including the potential regulator and signaling pathways of CD44. MATERIAL AND METHODS We searched the miRNA online database (www.mirdb.org) with the "seed sequence" located within the 3'-UTR of the target gene, and performed luciferase assay to test the miRNA/mRNA relationship. We also determined the expression of CD44 in the EGC and control samples. In addition, statistical analysis was used to explore the role of miR-328 as a biomarker to predict the recurrence after ECD. RESULTS We validated CD44 to be the direct gene via luciferase reporter assay system. We also established the negative regulatory relationship between miR-328 and CD44 via studying the relative luciferase activity at different concentrations of miR-328 mimics. We also conducted real-time PCR and Western blot analysis to study the mRNA and protein expression level of CD44 among different groups (recurrence-positive and recurrence-negative) or cells treated with different concentrations of miR-328 mimics/inhibitors, indicating the negative regulatory relationship between miR-328 and CD44. We also investigated the relative viability of EGC cells when transfected with miR-328 mimics (50 nM and 100 nM) and miR-328 inhibitors (100 nM) to validate miR-328 to be negatively interfering with the viability of EGC cells. miR-328 was also recognized as a potential biomarker to predict recurrence after ESD in EGC patients via analysis of the recurrence-free rate among different groups of EGC patients. CONCLUSIONS The expression level of miR-328 can function as a predictive biomarker of recurrence after ECD in patients with EGC via targeting CD44.

Effects of edge disorder on the stability of quantum oscillations in two-dimensional coupled systems.

This paper utilizes the theory of quantum diffusion to analyze the electron probability and spreading width of a wavepacket on each layer in a two-dimensional (2D) coupled system with edge disorder, aiming to clarify the effects of edge disorder on the stability of the electron periodic oscillations in 2D coupled systems. Using coupled 2D square lattices with edge disorder as an example, we show that, the electron probability and wavepacket spreading width exhibit periodic oscillations and damped oscillations, respectively, before and after the wavepacket reaches the boundary. Furthermore, these electron oscillations exhibit strong resistance against disorder perturbation with a longer decay time in the regime of large disorder, due to the combined influences of ordered and disordered site energies in the central and edge regions. Finally, we numerically verified the universality of the results through bilayer graphene, demonstrating that this anomalous quantum oscillatory behavior is independent of lattice geometry. Our findings are helpful in designing relevant quantum devices and understanding the influence of edge disorder on the stability of electron periodic oscillations in 2D coupled systems.

[Determination of hydrogen selenide in workplace air with atomic fluorescence].

OBJECTIVE: To determined hydrogen selenide in workplace air with atomic fluorescence. METHOD: Hydrogen selenide were sampled with 0.1 mol/L sodium hydroxide solution in multi-hole absorbing tubes. The sampled absorbing solution were digested with (9+1) nitric acid/perchloric acid. The selenide in sample were reduced by potassium borohydride in 5.0% hydrochloride solution and determined with atomic fluorescence. RESULTS: There was a good linearity (r=0.9999) over the concentration of 0-150 microg/L, The precision of low, middle and high concentration were 3.1%, 7.4% and 6.7%, respectively. The sample collection rate can reach 99%. CONCLUSION: The method was accurate and sensitive to detect hydrogen selenide in workplace air.

[The improvement of determination method of carbon disulfide in workplace air with gas chromatography].

OBJECTIVE: Establishment of determination method of carbon disulfide in charcoal tube with low toxicity solvents desorption-gas chromatography. METHODS: Four types of solvent with low toxicity are applied respectively as substitution of benzene to desorb the carbon disulfide in samples of charcoal tube. The signal strength and desorption efficiency of the detector are compared by using different solvents. RESULTS: Chloroform has been considered as the best alternative solvent of benzene. Carbon disulfide has a good linearity (R = 0.9997) over the concentration of 0 ∼ 54.7 µg/ml, detection limit can reach 0.2 µg/ml. When the sampling volume is 3.0 L, the minimum detectable concentration is 0.07 mg/m(3). CONCLUSION: With the use of chloroform, the health hazard to laboratory personnel and environment pollution as well as the costs of experiments are reduced.

[The optimization of 2-thiothiazolidine-4-carboxylic acid back extraction in urine and determination by HPLC].

OBJECTIVE: Establishment of determination method of 2-thiothiazolidine-4-carboxylic acid (TTCA) in urine with HPLC. METHODS: A volume of 0.5 ml hydrochloric acid (2 mol/L) and 0.5 ml pure water was added into 1 ml urine, and then extracted by 4 ml of diethyl ether by shaking for 2 min. Remove the water phase in a tube with plug and extract again, mix the two extraction diethyl ether together, take 4 ml by adding 2 ml borax-monopotassium phosphate buffer and shaking for 2 min to extract, then take the water phase to detect. A C(18) column and UV detector were used for separating and detecting. The wavelength was 273 nm, the flow rate was 1.0 ml/min, and the injection volume was 20 µl. RESULTS: TTCA has a good linearity (r = 0.9995) over the concentration of1 1 ∼ 10 µg and the minimum detectable concentration of TTCA in urine was 0.1 µg/ml. The within-day precision (RSD) were 8.4%, 3.0% and 1.7%, the between-day precision (RSD) were 11%, 3.8%, 1.9%, respectively. The extraction recovery were between 80% ∼ 102%. CONCLUSION: The method was accurate and sensitive to detect TTCA in urine.

[The rervising of biological exposure index for carbon disulfide exposuring].

OBJECTIVE: To study the biological exposure index of carbon disulfide in China. METHODS: High-performance liquid chromatography (HPLC) was used to detect the levels of 2-thiothiazolidine-4-carboxylic acid (TTCA) in the urine of the workers after working shift end, Gas chromatography was used to detect the concentrations of the carbon disulfide in the workplace air. The relationship between the urine TTCA levels and the concentrations of the carbon disulfide was analyzed, the biological exposure index and judgement result from PC-TWA were compared. RESULTS: The levels of TTCA in urine of workers occupationally exposed to carbon disulfide were closely and positively related with the concentrations of the carbon disulfide in the workplace air. The regression equation was Y = 0.265X - 0.165, The biological exposure index of carbon disulfide were calculated by regression equation according to occupational exposure limits of carbon disulfide in China. CONCLUSION: The biological exposure index of CS(2) in China might be revised for 1.2 mg/g Cr.

[Biological limit value for occupational exposure to N, N-dimethylacetamide].

OBJECTIVE: To establish Biological Limit Value (BLV) for N, N-dimethylacetamide (DMAC). METHOD: 201 workers in 3 spandex factories exposed to DMAC were recruited. Air samples were collected using personal air samplers, and urine samples from each works were collected at the end of shift at end of workweek. The urinary metabolite NMAC and air samples of DMAC were determined by gas chromatography (GC). Percentile and relative internal exposure (RIE) were analyzed and proposed a BLV for DMAC. RESULTS: The number of workers who exposure to DMAC below OELs were 133 (66.2%) among 201 workers monitored. Geometric mean (range) concentration of DMAC in air was 19.4 (0.40 ∼ 300.12) mg/m(3), and that of NMAC in urine was 23.7 (1.30 ∼ 189.42) mg/g Cr. A linear correlation was found between the personal air DMAC and creatinine-adjusted NMAC levels in urine collected at the end of shift at end of workweek (F = 188.872, R(2) = 0.487,P < 0.001). The relationship can be described by the equation Log (NMAC mg/g Cr) = 0.685 + 0.455 log (DMAC mg/m(3)). According to the equation the current China OELs value of 20 mg/m(3) would lead to a mean NMAC concentration of 18.92 mg/g Cr. The 90th percentile biomonitoring result below 20 mg/m(3) 8-hour TWA is 23.9 mg MMAC mg/g Cr, and that of NMAC in urine calculated by relative internal exposure (RIE) was 19.0 mg/g Cr. CONCLUSION: A BLV of 20 mg/g Cr NMAC in urine at the end of shift at end of workweek for DMAC was recommend by reference to official values from other countries.

[The interventions effect-assessment of the workers exposed to N, N-dimethylformamide by percutaneous in a synthetic leather factory].

OBJECTIVE: To assess the effects of interventions on synthetic leather workers exposed to N, N-dimethylformamide (DMF) by skin. METHODS: Twenty-six workers exposed to DMF were recruited. The level of DMF in ambient or handwash solution and N-methylformamide (NMF) in end-shift urine samples were detected before interventions and after interventions for six months. RESULTS: After interventions the levels of DMF in ambient reduced 52.7% from (63.27 +/- 52.67) mg/m3 to (29.95 +/- 23.79) mg/m3. The levels of NMF in urine samples reduced 17.9% from (2.07 +/- 0.32) mg/g Cr to (1.70 +/- 0.29) mg/g Cr (P < 0.01). The mean level of DMF in handwash solution reduced 53.4% from 0.88 +/- 0.40 mg to 0.41 +/- 0.81 mg. CONCLUSION: This study showed that the multi-intervention measures (engineering control, personal protection and health promotion) should be used for the synthetic leather workers occupationally exposed to DMF.

[The impact of N, N-dimethylacetamide on the health of workers].

OBJECTIVE: To explore the hepatic toxicity and the exposure biomarkers of N, N-Dimethylacetamide. METHODS: One hundred forty five objects were chosen by stratified random sampling method. The investigation was performed using questionnaire and physical examination. The air concentrations of DMAC in the workshops were monitored. The urine samples were collected and analyzed after work everyday or at the weekend. The correlation between the air concentrations of DMAC in the workshops and the concentrations of urinary NMAC wee analyzed by regression. RESULTS: The air concentration of DMAC in the spinning workshop was higher than others. The morbidity of abnormal hepatic function was 12.4%, 61.1% of workers with abnormal hepatic function appeared in one year after exposure to DMAC in the workshops ( r=0.44, P<0.01). CONCLUSION: The abnormal heptic function was found in workers exposed to DMAC for short period. The concentration of urinary NMAC can serve as the exposure biomarker of DMAC.

Secondary Cleft Lip Induced by Congenital Hemangioma: Report of a Rare Case.

Infant hemangioma is a relatively rare congenital disease. Ulcer and infection may occur in some cases. A few cases may develop into deformity and defect. Herein, we present a case of secondary cleft lip induced by congenital hemangioma and also our sequential treatment.

Bushen Qiangjin capsule inhibits the Wnt/α-catenin pathway to ameliorate papain-induced knee osteoarthritis in rat.

OBJECTIVE: To evaluate the molecular mechanism underlying the beneficial effect of Bushen Qiangjin capsule (BSQJ), a Traditional Chinese Medicine, on knee osteoarthritis (KOA). METHODS: In the present study, 32 female Sprague-Dawley rats were randomly divided into four groups: control, KOA, high-dose BSQJ (H-BSQJ), and low-dose BSQJ (L-BSQJ). After successfully establishing the KOA model by intra-articular injection of papain, H-BSQJ and L-BSQJ groups were intragastrically administered 0.243 and 0.122 g/kg BSQJ, respectively, daily for 6 weeks. At the end of the experiment, knee articular cartilage tissues of rats were collected for evaluation by hematoxylin and eosin staining, Safranin O-Fast Green staining, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. Serum interleukin-1α and tumor necrosis factor-α levels of rats were detected with an enzyme-linked immunosorbent assay method. Gene expression of Wnt-4, α-catenin, Frizzled-2, glycogen synthase kinase-3ß (GSK-3ß), cysteinyl aspartate-specific proteinases 3 and 9 (caspases 3 and 9), collagen type II alpha 1 (Col2a1), and matrix metalloproteinases 1 and 13 (MMP-1 and MMP-3) of rat knee articular cartilage was quantified by reverse transcription-quantitative polymerase chain reaction analysis. Wnt-4, α-catenin, Frizzled-2, GSK-3ß, cleaved caspase-3, and cleaved caspase-9 protein expression in rat knee articular cartilage was determined by western blot analysis. RESULTS: BSQJ obviously reduced pathological damage and matrix degradation of articular cartilage in KOA rats. Compared with the KOA group, H-BSQJ rats exhibited downregulated mRNA and protein expression of Wnt-4, ß-catenin, Frizzled-2,and caspase-3, as well as upregulated mRNA and protein expression of GSK-3α. In addition, H-BSQJ significantly increased mRNA expression of Col2a1 and decreased mRNA expression of MMP-1 and MMP-13. CONCLUSION: BSQJ exerted a beneficial effect on KOA by a mechanism involving downregulation of the Wnt/α-catenin pathway, which inhibited both cartilage extracellular matrix degradation and chondrocyte apoptosis to ameliorate KOA in rats.

Clinical assessment of synbiotics for treating chronic kidney disease: A protocol for systematic review and meta-analysis.

BACKGROUND: This research aims to evaluate the efficacy and safety of synbiotics for treating chronic kidney disease. METHODS: Related articles written in English were sourced from EMBASE, PubMed, Cochrane Library, and China National Knowledge Infrastructure. These articles were used in the evaluation of the effect of synbiotics for treating chronic kidney disease. The extent of the relationship was assessed by calculating the pooled risk ratio, mean differences or standardized mean difference along with the equivalent 95% confidence intervals. The risk of bias introduced through each study was considered by adopting the Cochrane Risk of Bias Tool. Suitable statistical research methods were utilized for the synthesis of the data. The RevMan 5.3 software was used to conduct all statistical analysis. RESULTS: The final results of the current study is due to be included in a peer-reviewed journal. CONCLUSION: The final remarks of the current study will be useful evidence for determining whether synbiotics is an effective and safe therapeutic method for treating chronic kidney disease. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/UASF4 (https://osf.io/uasf4/).

[DMF-induced adult hepatocyte apoptosis and its effects on expression of Bcl-2, Bax and Caspase-3].

OBJECTIVE: To investigate the apoptosis-inducing effect of DMF on the human liver cells (HL-7702) in vitro. METHODS: Liver cells were exposed to different concentrations of DMF (0, 50, 100, 150, 200 mmol/L) for 12 hours. Apoptotic rate, the expression of Bax, Bcl-2 and Caspase-3 in liver cells were measured by FCM and western blotting respectively. RESULTS: The increase in apoptotic rate of hepatocytes in concentration-manner was shown after DMF treatment for 12 h. After treatment the expression of Bcl-2 was decreased steadily and lower than the control group (P < 0.01), the expression of Bax showed no significant difference among the groups of different dosage by one-factor analysis of variance (P > 0.05), as the increase of the dosage of DMF. The ratio of Bcl-2/Bax dropped with the dosage of DMF increasing, and the ratio in 200 mmol/L of DMF was significantly lower than that of the control (P < 0.01). The new lands of procaspase-3 in 150, 200 mmol/L were observed, which demonstrated that there was active caspase-3. CONCLUSION: DMF can induce apoptosis of cultured adult normal hepatocytes in vitro, and the mechanism might be related to the decrease of Bcl-2/Bax and the cleavage of Caspase-3.

Oral respiratory syncytial virus (RSV) DNA vaccine expressing RSV F protein delivered by attenuated Salmonella typhimurium.

Human respiratory syncytial virus (RSV) is a major viral pathogen of the lower respiratory tract of infants and young children worldwide. No effective prevention measure is available. Attenuated Salmonella strains expressing heterologous antigens can be delivered by the oral route, triggering efficient antigen-specific humoral, cellular, and mucosal immunity. In this study, we orally administered attenuated Salmonella strain SL7207, carrying the plasmid pcDNA3.1/F expressing the RSV F gene, to BALB/c mice and showed significant elevations of serum anti-RSV IgG and bronchoalveolar lavage secretory IgA as compared with the control group carrying empty plasmid (p<0.001). The ratio of IgG1 and IgG2a was 0.96. The experimental group also showed a stronger cytotoxic T cell response (p<0.01 at effector:target ratios of 100:1 and 50:1) and a higher stimulation index value of T cell proliferation (p<0.05) than the respective control group. RSV titers in the lung homogenates of the experimental group on day 3 and day 5 postchallenge were lower than in the control group (p<0.05). Histopathological analysis showed obvious differences in infiltration of inflammatory cells and pulmonary alveolar wall thickness (p<0.01) between the two groups. In summary, our results demonstrate the potential of orally administered SL7207-based DNA vaccines against RSV infection.

[Effect of Hedgehog Signaling Pathway Abnormality on Chemothe-rapeutic Resistance of Multiple Myeloma].

OBJECTIVE: To study the correlation between the excessive activation of Hedgehog signal and the drug resistance of multiple myeloma. METHODS: The resistant cell line RPMI 8226/R of multiple myeloma was established by an ascending concentration gradient method. The experiment consisted of 4 groups: RPMI8226/R, RPMI8226/S, GANT61+RPMI8226/R and GANT61+RPMI8226/S. The CCK-8 (cell counting kit-8) assay was used to detect the cell proliferation inhibition rate in 4 groups; the RT-PCR was used to detect the expression of Gli1, Gli2, Shh, Ihh, Smo and Sufu in the RPMI8226/S and RPMI8226/R cells. The Western blot was used to detect the expression of the resistant protein Cyclin D1, P21 and BCL-2 and MDR-related signaling pathways protein p-Akt, p-MAPK and STAT3 in the RPMI8226/S and RPMI8226/R cells. After adding different concentration of GANT61, the Western blot was used to detect the expression of Gli2 in RPMI8226/R and RPMI8226/S cells. RESULTS: The expression of Shh, Ihh, Smo Gli2 was enhanced significantly in the RPMI8226/R cells, but the expression of Sufu inhibitor was reduced, the expression level of related protein in Hedgehog signaling pathway was significanly higher in RPMI8226/R than that in RPMI8226/S. After theatment of GANT61 in vitro, the expression level of Gli2 in multiple myelom cells obviously decreased, the decreasing effect of GANT61 on Gli2 expression in RPMI8226/R cells was more significant than that in RPMI8226/S cells. The sensitivity of RPMI8226/R cells to DOX after treatment with GANT61 (IC50) was risen from 7.11±0.061 µmol/L to 0.99±0.053 µmol/L, the corresponding cell resistance index decreased from 5.51 to 1.69. CONCLUSION: the activation of Hedgehog signaling pathway is closely related with the resistance of multiple myeloma cells, and GANT61 can block the Hedgehog signaling pathway, thus Hedgehog signaling may be used as a new target for multiple myeloma treatment.

Effects of dexmedetomidine versus midazolam for premedication in paediatric anaesthesia with sevoflurane: A meta-analysis.

Background Dexmedetomidine (DEX), an α2-adrenergic receptor agonist, produces ideal sedation and early postoperative recovery for premedication in paediatric surgery, reducing preoperative anxiety and facilitating smooth induction of anaesthesia. We performed a meta-analysis to compare the effects of DEX and midazolam (MDZ) in paediatric anaesthesia with sevoflurane. Methods PubMed, Ovid, Web of Science, and Public Health Management Corporation were searched through December 2016 for randomized controlled trials (RCTs) that compared DEX and MDZ in children undergoing sevoflurane anaesthesia. The risk ratio (RR) with 95% incidence interval (95%CI) was used for dichotomous variables. Results Twelve RCTs involving 422 patients in the DEX group and 448 patients in the MDZ group were included. Patients in the DEX group had a significantly lower incidence of unsatisfactory sedation (RR [95%CI] = 0.71 [0.57-0.89]), unsatisfactory parental separation (RR [95%CI] = 0.56 [0.35-0.87]), and rescue analgesia (RR [95%CI] = 0.52 [0.35-0.77]) than patients in the MDZ group. However, both groups had a similar incidence of unsatisfactory mask acceptance, emergence agitation, and postoperative nausea and vomiting. Conclusion Compared with MDZ, DEX is beneficial in paediatric anaesthesia with sevoflurane because of its lower incidence of unsatisfactory sedation, parental separation, and rescue analgesia.

[Influences of biochar and nitrogen fertilizer on soil nematode assemblage of upland red soil].

The use of biochar as soil remediation amendment has received more and more concerns, but little attention has been paid to its effect on soil fauna. Based on the field experiment in an upland red soil, we studied the influences of different application rates of biochar (0, 10, 20, 30, 40 t · hm⁻²) and nitrogen fertilizer (60, 90, 120 kg N · hm⁻²) on soil basic properties and nematode assemblages during drought and wet periods. Our results showed that the biochar amendment significantly affect soil moisture and pH regardless of drought or wet period. With the increasing of biochar application, soil pH significantly increased, while soil moisture increased first and then decreased. Soil microbial properties (microbial biomass C, microbial biomass N, microbial biomass C/N, basal respiration) were also significantly affected by the application of biochar and N fertilizer. Low doses of biochar could stimulate the microbial activity, while high doses depressed microbial activity. For example, averaged across different N application rates, biochar amendment at less than 30 t · hm⁻² could increase microbial activity in the drought and wet periods. Besides, the effects of biochar also depended on wet or drought period. When the biochar application rate higher than 30 t · hm⁻², the microbial biomass C was significantly higher in the drought period than the control, but no differences were observed in the wet period. On the contrary, microbial biomass N showed a reverse pattern. Dissolved organic matter and mineral N were affected by biochar and N fertilizer significantly in the drought period, however, in the wet period they were only affected by N fertilizer rather than biochar. There was significant interaction between biochar and N fertilizer on soil nematode abundance and nematode trophic composition independent of sampling period. Combined high doses of both biochar and N fertilization promoted soil nematode abundance. Moreover, the biochar amendment increased the proportion of fungivores especially in the drought period, suggesting the biochar was the preferred fungal energy channel in comparison to soil without biochar addition. In summary, complex patterns occurred not only due to the application rate of biochar as well as their interactions with N fertilization but also due to the drought and wet periods. It is, therefore, necessary to consider different ecological factors when evaluating the effects of biochar in future.

Neuroprotective effects of glial cell line-derived neurotrophic factor mediated by an adeno-associated virus vector in a transgenic animal model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a relentlessly progressive lethal disease that involves selective annihilation of motoneurons. Glial cell line-derived neurotrophic factor (GDNF) is proposed to be a promising therapeutic agent for ALS and other motor neuron diseases. Because adeno-associated virus (AAV) has been developed as an attractive gene delivery system with proven safety, we explored the therapeutic efficacy of intramuscular delivery of the GDNF gene mediated by an AAV vector (AAV-GDNF) in the G93A mouse model of ALS. We show here that AAV-GDNF leads to substantial and long-lasting expression of transgenic GDNF in a large number of myofibers with its accumulation at the sites of neuromuscular junctions. Detection of GDNF labeled with FLAG in the anterior horn neurons, but not beta-galactosidase expressed as a control, indicates that most of the transgenic GDNF observed there is retrogradely transported GDNF protein from the transduced muscles. This transgenic GDNF prevents motoneurons from their degeneration, preserves their axons innervating the muscle, and inhibits the treated-muscle atrophy. Furthermore, four-limb injection of AAV-GDNF postpones the disease onset, delays the progression of the motor dysfunction, and prolongs the life span in the treated ALS mice. Our finding thus indicates that AAV-mediated GDNF delivery to the muscle is a promising means of gene therapy for ALS.