The Role of NF mRNA and Calpain in NF Reduction of Acrylamide Neuropathy.

The purpose of this study was to study the role of neurofilament (NF) mRNA and calpain in NF reduction of acrylamide (ACR) neuropathy. Male Wistar adult rats were injected i.p. every other day with ACR (20 mg/kg·bW or 40 mg/kg·bW) for 8 weeks. NF mRNA expression was detected using RT-PCR and the calpain concentration was determined using western blot analysis. The NF mRNA expression significantly decreased while the level of m-calpain and µ-calpain significantly increased in two ACR-treated rats groups regardless of the ACR dose. The light NF (NF-L) protein expression was significantly correlated with NF-L mRNA expression. Combined with previous data, the concentrations of three NF subunits were negatively correlated with the calpain levels. These findings suggest that NF-L mRNA and calpain mediated the reduction in NF of ACR neuropathy.

[Analyzing the factors of influencing the musculoskeletal disorders of greenhouse vegetable farmers].

OBJECTIVE: To study the prevalence of musculoskeletal disorders (MSDs) among greenhouse vegetable farmers and to explore the risk factors of MSDs. METHODS: A household questionnaire survey was conducted to investigate 203 greenhouse vegetable farmers and 127 non-greenhouse vegetable farmers in February, 2011. RESULTS: The one-year prevalence rates of MSDs were 70.0% and 33.9% among greenhouse vegetable farmers and non-greenhouse vegetable farmers, respectively. The three main positions of MSDs in greenhouse farmers were low back, knee (s) and shoulder (s). Age, working years, body weight and usage of rolling machine were statistically associated with MSDs of greenhouse farmers, ORadj values were 1.17, 1.82, 1.08 and 0.07, respectively. CONCLUSIONS: The prevalence of MSDs is high in greenhouse workers. Low back pain, knee (s), and shoulder (s) disorders are the main disorders. Age, working years, body weight and usage of rolling machine are main risk factors for the development of MSDs in greenhouse farmers.

First identification of methicillin-resistant Staphylococcus aureus MLST types ST5 and ST45 and SCCmec types IV and Vt by multiplex PCR during an outbreak in a respiratory care ward in central Taiwan.

We used molecular typing methods to investigate an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) infections in a respiratory care ward in Taiwan. From March to June 2006, the incidence of MRSA infection increased 3.75-fold. The overall carrier rates among the health care workers (HCWs) were 31.3% (total S. aureus), 16.4% (MRSA), and 14.9% (methicillin-sensitive SA, MSSA). Pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), antibiograms derived from susceptibility testing of MRSA isolates, and multiplex polymerase chain reaction (PCR) provided strong epidemiologic and microbiologic evidence that the outbreak of MRSA infections at our hospital was linked to the same PFGE pulsotype A SCCmec type II, pvl-negative, MLST ST5 strain of MRSA isolated from seven HCWs and five patients. The outbreak was controlled by application of topical fucidin ointment to the anterior nares in all colonized HCWs. Multiplex PCR combined with PFGE and MLST is a feasible method for outbreak investigations in routine clinical laboratories.

Antimicrobial resistance of bacterial isolates from respiratory care wards in Taiwan: a horizontal surveillance study comparison of the characteristics of nosocomial infection and antimicrobial-resistant bacteria in adult Intensive Care Units and two respiratory care facilities for mechanically ventilated patients at a tertiary care centre in Taiwan.

The objectives of this study were to compare the incidence of nosocomial infections (NIs) and the distribution of resistant nosocomial pathogens in adult Intensive Care Units (ICUs) and two respiratory care facilities for prolonged mechanically ventilated patients [i.e. the respiratory care centre (RCC) and the respiratory care ward (RCW)] in a 1100-bed tertiary care hospital in Taiwan from 2003 to 2006. The overall incidences of NI for adult ICUs, the RCC and the RCW were 14.0, 10.3 and 5.0 per 1000 patient-days, respectively. Urinary tract infections, bloodstream infections and pneumonias occurred most frequently. The most common reported microorganisms in adult ICUs were non-fermentative Gram-negative bacilli (NFGNB) (33.0%), Enterobacteriaceae (26.5%), Candida spp. (18.2%), Staphylococcus aureus (8.9%), coagulase-negative staphylococci (CoNS) (4.9%) and enterococci (4.5%). In comparison, RCW patients had a higher proportion of NIs caused by S. aureus [odds ratio (OR)=1.9], enterococci (OR=2.2) and Enterobacteriaceae (OR=2.2), but a lower proportion of CoNS (OR=0.3), NFGNB (OR=0.5) and Candida spp. (OR=0.2). RCW patients had higher incidence rates of methicillin-resistant S. aureus (OR=4.91) and extended-spectrum ß-lactamase-producing Enterobacteriaceae (OR=4.06) than ICU patients. Further study is needed to delineate the mechanisms responsible for the differences in resistance profile amongst pathogens associated with nosocomial infection in ICUs, RCCs and RCWs.

2,5-hexanedione (HD) treatment alters calmodulin, Ca2+/calmodulin-dependent protein kinase II, and protein kinase C in rats' nerve tissues.

Calcium-dependent mechanisms, particularly those mediated by Ca(2+)/calmodulin (CaM)-dependent protein kinase II (CaMKII), have been implicated in neurotoxicant-induced neuropathy. However, it is unknown whether similar mechanisms exist in 2,5-hexanedione (HD)-induced neuropathy. For that, we investigated the changes of CaM, CaMKII, protein kinase C (PKC) and polymerization ratios (PRs) of NF-L, NF-M and NF-H in cerebral cortex (CC, including total cortex and some gray), spinal cord (SC) and sciatic nerve (SN) of rats treated with HD at a dosage of 1.75 or 3.50 mmol/kg for 8 weeks (five times per week). The results showed that CaM contents in CC, SC and SN were significantly increased, which indicated elevation of Ca(2+) concentrations in nerve tissues. CaMKII contents and activities were also increased in CC and were positively correlated with gait abnormality, but it could not be found in SC and SN. The increases of PKC contents and activities were also observed in SN and were positively correlated with gait abnormality. Except for that of NF-M in CC, the PRs of NF-L, NF-M and NF-H were also elevated in nerve tissues, which was consistent with the activation of protein kinases. The results suggested that CaMKII might be partly (in CC but not in SC and SN) involved in HD-induced neuropathy. CaMKII and PKC might mediate the HD neurotoxicity by altering the NF phosphorylation status and PRs.

Effects of acrylamide on the nervous tissue antioxidant system and sciatic nerve electrophysiology in the rat.

To investigate the time-dependent effects of acrylamide (ACR) on the antioxidative status in rat nerve tissues, adult male Wistar rats were given ACR (40 mg/kg, i.p., 3 times/week) for 2, 4, 6 and 10 weeks, respectively. The time-dependent changes of the lipid peroxidation (malondialdehyde, MDA) and antioxidative status (glutathione, GSH; glutathione peroxidase, GSH-Px; glutathione reductase, GR; superoxide dismutase, SOD and anti-reactive oxygen species, anti-ROS) in nerve tissues were investigated. The electrophysiology indices (nerve conduction velocity, NCV; compound action potential duration, CAPD; compound action potential amplitude, CAPA; compound action potential latency, CAPL) in the sciatic nerve were determined using BL-420E Biologic Function Determining System. The results showed that MDA levels increased significantly (P < 0.05) in nerve tissues, while GSH levels markedly decreased (P < 0.05) in a time-dependent manner. SOD activity (in the spinal cord and sciatic nerve) and GR activity (in the sciatic nerve) increased significantly after 4 weeks ACR treatment (P < 0.01), but then decreased (P < 0.05). The anti-ROS activity in the sciatic nerve was markedly decreased at the end of week 6 and 10 (P < 0.01). The above indices changed most in the sciatic nerve. The levels of GSH, MDA and anti-ROS in rat sciatic nerve were in high correlation (P < 0.05, |r| > 0.80) with the electrophysiology indices according to the exposure time. Thus, ACR-induced neurotoxicity may be associated with the enhancement of lipid peroxidation and reduction of the antioxidative capacity. Depletion of neural GSH level might be one of the primary events in ACR-induced neuropathy.

Malondialdehyde and catalase as the serum biomarkers of allyl chloride-induced toxic neuropathy.

Chronic exposure to allyl chloride (AC) is known to produce a central-peripheral distal axonopathy. To access the biomarker of exposure and elucidate the mechanism of neuropathy induced by AC, we performed a longitudinal observational study of malondialdehyde (MDA), anti-reactive oxygen species (anti-ROS), glutathione (GSH), catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD) in rats serum and sciatic nerve after 0, 3, 6, 9 and 12 weeks of AC administration. AC was administrated to Wistar rats by gavage at a single dosage of 200 mg/kg/per dose (three times per week). Rats were sacrificed after 0, 3, 6, 9 and 12 weeks of AC treatment, serum and sciatic nerves were quickly collected at 4 degrees C. The results showed that MDA levels in serum (115.4 and 126.2%) and sciatic nerve (130.5 and 145.3%) significantly increased (p<0.05) on 3rd week of AC treatment and at gait score of 2, and further changes of MDA levels were observed after 6, 9 and 12 weeks and at gait score of 3 and 4. While a decrease (p<0.05) in the activities of CAT on 6th week of AC intoxication and at gait score of 2 was observed in serum (81.2 and 72.8%) and sciatic nerve (71.7 and 70.7%). The other antioxidants also decreased in serum and sciatic nerve after 3, 6 and 9, 12 weeks' intoxication and at gait score of 2, 3 and 4. Significant (p<0.05) positive correlations were observed between serum and sciatic nerve in MDA levels (r=0.9162 and 0.9551, respectively) and CAT (r=0.9410 and 0.9557, respectively) activities as time went on and symptoms developed. Thus, AC intoxication was associated with elevation of lipid peroxidation and reduction of antioxidative status, and the time dependent changes of these indexes in Wistar rats' serum and sciatic nerve occurred. The misbalance of lipid peroxidation and antioxidation status might be one of mechanisms of toxic neuropathy induced by AC. MDA and CAT could be served as the biomarkers of AC exposure to afford the early diagnosis of AC-induced toxic neuropathy.

Expression changes of neurofilament subunits in the central nervous system of hens treated with tri-ortho-cresyl phosphate (TOCP).

Tri-ortho-cresyl phosphate (TOCP) could induce degeneration of long, large diameter axons within the central and peripheral nervous system of susceptible species including human being and hens, which is referred to as organophosphorus-ester induced delayed neuropathy (OPIDN). The mechanisms involved are not understood. Neuropathologic observations suggested that neurofilament subunits (NFs) could be a main target of TOCP in the peripheral nervous system. Our previous study also showed that NFs in protein levels significantly decreased in sciatic nerves of hens treated with TOCP. In this study, to determine whether the decrement of NFs proteins in sciatic nerves was due to reductions in NF gene expression or protein degradation, hens were treated with a single dose of 750 mg/kg body weight TOCP by gavage, and sacrificed on 21 day post-exposure. Cerebral cortexes and spinal cords were sampled. Transcriptional changes of NFs including high molecular weight neurofilament (NF-H), middle molecular weight neurofilament (NF-M), low molecular weight neurofilament (NF-L), and glyceraldehydes-3-phoaphate dehydrogenase (GAPDH) as inner inference in cerebral cortexes and spinal cords were analyzed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). Results showed that all of three NFs mRNA in cerebral cortexes down-regulated significantly. However, in spinal cords, there was only NF-M decreased, both of NF-H and NF-L kept unaffected. The protein levels of NFs in pellet and supernatant fractions of cerebral cortexes and spinal cords were also determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. We noticed that all NFs protein declined in pellet of cerebral cortexes, but NF-M reduction was not significant compared with that of control hens. NF-H and NF-M proteins in supernatant of cerebral cortexes exhibited significant increase, while NF-L level showed remarkable decline. In spinal cords, apart from NF-L in pellet were significantly increased, both of NF-H and NF-M in pellet and supernatant, as well as NF-L in supernatant fractions were manifested dramatic reduction compared with the pattern of control. The quantitative analyses revealed that the change magnitude in protein levels was much greater than that in mRNA levels in hens' central nervous system after TOCP administration. These findings suggest that the NFs disturbance in protein levels is closely associated with the decreases in sciatic nerves observed in our previous work after TOCP exposure, rather than that in mRNA levels, and the NFs alterations in protein levels may be one of the responsible factors for the OPIDN.

Effect of subchronic exposure to acrylamide induced on the expression of bcl-2, bax and caspase-3 in the rat nervous system.

Occupational exposure and experimental intoxication with acrylamide (ACR) produce a neuropathy characterized by nerve degeneration. We hypothesize that ACR differentially affects the expression of bcl-2, bax and caspase-3 in the central nervous system (CNS) and the peripheral nervous system (PNS) tissue. Male adult Wistar rats were given ACR (20, 40 mg/kg i.p. 3 days/week) for 8 weeks. Samples of the cerebral cortex, cerebellum, spinal cord and sciatic nerves were collected and examined for bcl-2, bax and caspase-3 expression using Western blotting. Subchronic exposure to ACR reduced cortical bcl-2 expression in the low dose, increased it in the high dose; the change of bcl-2 expression in the spinal cord and cerebellum followed the same pattern as that described in the cerebral cortex; there was no significant change in the expression of bax in the cerebral cortex and the spinal cord, however, in the cerebellum the change of bax expression and bcl-2 expression is just the reverse. Thus, the bcl-2/bax ratio of the CNS tissue was affected by exposure to ACR, it decreased in the low dose group and increased in the high group. Compared to control, densitometric analysis showed that in the sciatic nerves the expression of bcl-2 and bax expression was markedly increased following ACR administration. The expression of inactive isoforms (32 kDa) of caspase-3 was not altered in the cortices of ACR-treated rats, but increased in their spinal cords and sciatic nerves. Thus, subchronic exposure to ACR affected the expression of death-related proteins in the CNS and PNS tissue, which indicate there is the early molecular regulatory mechanism of apoptosis in the neuropathy induced by ACR.

Allyl chloride-induced time dependent changes of lipid peroxidation in rat nerve tissue.

To accurately know the time-dependent changes of the lipid peroxidation and antioxidative status for elucidating the mechanism of neuropathy induced by allyl chloride (AC), the malondialdehyde (MDA), anti-reactive oxygen species (anti-ROS), glutathione (GSH), catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD) were investigated in cerebrum, spinal cord and sciatic nerve of rats after 0, 3, 6, 9, 12 weeks of AC administration. AC was administrated to Wistar rats by gavage at a single dosage of 200 mg/kg/per dose (three times per week). Rats were sacrificed after 0, 3, 6, 9, 12 weeks of treatment, and cerebrum, spinal cord, sciatic nerves were dissected, homogenized and used for the determination of lipid peroxidation and antioxidative status. The results showed that MDA in cerebrum (112.4%) and sciatic nerve (113.1%) significantly increased (P<0.05) on third week of AC treatment and at gait score of 2, and further changes of MDA were observed after 6, 9, 12 weeks and at gait score of 3, 4. While a decrease (P<0.05) in the activities of GSH, CAT, GPx and SOD after 6, 9, 12 weeks intoxication and at gait score of 2, 3, 4 were observed in cerebrum, spinal cord and sciatic nerve. Anti-ROS activities also decreased in all three nerve tissues after 3, 6, 9, 12 weeks intoxication and at gait score of 2, 3, 4. Thus, AC intoxication was associated with elevation of lipid peroxidation and reduction of antioxidative status, and the time-dependent changes of these indexes in Wistar rats nerve tissues occurred. Sciatic nerve was the main target tissue and MDA was most sensitive among all indexes. The changes of lipid peroxidation and antioxidative status might be related to the degradation of nerve fiber and served as one of mechanisms of toxic neuropathy induced by AC.

2,5-Hexanedione induced reduction in protein content and mRNA expression of neurofilament in rat cerebral cortex.

Exposure chronically to n-hexane produces central-peripheral axonopathy mediated by 2,5-hexanedione (HD). Studies have shown neurofilament (NF) subunit proteins are decreased substantially in cerebral cortices, optic axons, spinal cords, and sciatic nerves from HD-exposed rats. To deeply investigate the alterations in NF contents in HD neuropathy, the relative levels of NF-L, NF-M, and NF-H in rat cerebral cortex were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. HD was administrated to Wistar rats by intraperitoneal injection at dosage of 200 or 400mg/kg. Rats were sacrificed after 6 weeks of treatment, and cerebral cortices were dissected, homogenized, and used for the determination of NF subunit proteins. The results, except for supernatant NF-L and NF-M that could not be assayed, showed HD intoxication resulted in significant decreases by 32-67% (P<0.01) in NF subunits in both of the pellet and supernatant fractions of cerebral cortex homogenate. As an initial investigation to determine how such changes in NF proteins might occur, the gene expression of NF-L, NF-M, and NF-H subunit mRNA was quantified using reverse transcription-polymerase chain reaction (RT-PCR). Statistical analysis revealed that HD exposure caused a significant reduction in the expression of NF-L and NF-H gene (P<0.05 or P<0.01), while the levels of NF-M mRNA kept unaffected (P>0.05). These suggest that the observed reduction in NF gene expression might be related to diminished levels of subunit proteins, while the actual contribution might be uncertain. The functional significance of the reduced protein contents and the regulation of gene expression remain to be determined.