Volume 5 (2011): Issue 3 (June 2011)

Background: Envenoming by snakebite is an important public health problem in rural tropics. Venomous snake families such as Elapidae and Viperidae frequently produce severe poisoning. Anti-venoms are not available for all venomous snakes of Thailand and there is need for more development in this field.

Objective: We characterized the important venomous snakes’ distribution of Thailand.

Method:Venomous snake species are described in details including their identification, range, and extraterritorial distribution.

Result: Eighteen snake species of the family Elapidae are summarized in their characteristics and distribution. There are three species of Naja, one species of Ophiophagus, three species of Bungarus, four species of Calliophis, one species of Sinomicrurus, two species of Laticauda, and four species of subfamily Hydrophiinae. Fifteen snake species of the family Viperidae consisting of one species of subfamily Viperinae and fourteen species of subfamily Crotalinae are also discussed.

Conclusion: All these snakes are venomous and their venom is potentially fatal since birth.

Background: We have previously reported that serum zinc (Zn) and copper (Cu) are affected by a number of factors. In the current investigation we have investigated the association between serum Zn and Cu concentrations and socio-economic factors in an Iranian population.

Materials and methods: A Persian sample population (n = 2233; n = 1106 (49.5%) males and n = 1127 (50.5%) females) was recruited by cluster-stratified sampling. Individuals were aged 15-65 years, and included urban and rural residents of the Great Khorasan province, Iran. Anthropometric measurements, serum Zn and Cu analysis and socio-economic status were determined using standard protocols.

Results: The mean serum Cu and Zn concentrations for the whole group were 14.7±3.3 μmol/L (range 4.5-28.4 μmol/L), and 11.7±1.9 μmol/L (range 3.6-28.3 μmol/L) respectively, and the mean serum Zn:Cu ratio for the group was 0.83±0.2. The highest mean copper concentrations were found in the age range 50-59 years (p < 0.01). The total population of urban residents had higher serum zinc (p <0.01) and lower serum copper concentrations (p <0.05) than rural residents. Poorly educated male subjects had significantly higher serum concentrations of copper than males in the other subgroups (p <0.001). Serum Cu and Zn:Cu ratio were associated with height and body mass indices (p <0.01).

Conclusion: Low serum zinc and copper appears to be common in Persian individuals. Urbanization and also educational attainment may contribute to changes in serum levels of Cu and Zn. This is probably related to lifestyle habits.

Background: In the kidney, angiotensin II (ANG II) and nitric oxide (NO) can stimulate each other. Unilateral ureteral obstruction (UUO) activates both substances, where ANG II is stimulated first and NO is augmented later.

Objective: Investigate the role of ANG II on renal nitric oxide synthase (NOS) protein expression in UUO.

Methods: Male Wistar rats were divided into sham and UUO. The UUO rats were treated separately with water, angiotensin converting enzyme inhibitor (ACEI), or angiotensin receptor type 1 blocker (ARB) for one day before UUO and continuously for one or seven days after the operation. The endothelial NOS (eNOS) and inducible NOS (iNOS) protein expressions were examined in histology.

Results: By immunohistochemistry, renal eNOS protein expression in the sham group showed staining in glomerulus and tubular epithelial cells in the cortex and medulla. UUO for one or seven days increased eNOS protein expression. ACEI or ARB reduced the heightened expression caused by UUO in 1-day group. However, in 7-day group, the elevated expression was maintained in the cortex, but was further increased in the medulla after ACEI or ARB administration. Both 1-day and 7-day UUO, with or without angiotensin blockade agents, caused no change in iNOS protein expression. One-day UUO resulted in mild tubular dilatation and cell infiltration. ACEI or ARB could attenuate structural alterations. The 7-day UUO rats demonstrated progressively morphological changes. ACEI was more effective than ARB in reducing tissue destruction.

Conclusion: In UUO, angiotensin blockade could attenuate renal eNOS protein expression in 1-day UUO group but not in 7-day UUO animals. The inhibition of angiotensin system ameliorates nephropathy induced by UUO.

Background: Curcumin (CUR) and tetrahydrocurcumin (THC) inhibits tumor angiogenesis. It is suggested that tumor progress may be related to the pathway of extracellular signal-regulated kinase 1/2 (ERK1/2) and serine/threonine kinase AKT, but the mechanism remains unclear.

Objective: Investigate the effects of CUR and THC on the expression of ERK1/2 and AKT in hepatocellular carcinoma (HepG2)-induced tumors in nude mice.

Methods: The curcuminoid mixture was obtained from the rhizomes of Curcuma longa, which was subjected to silica gel column chromatography to afford CUR as the major constituent. THC was prepared by hydrogenation of curcumin with palladium on charcoal as a catalyst. HepG2-implanted nude mice model was used to study of angiogenesis and tumor progression. Expressions of phospho-ERK1/2 (p-ERK1/2) and phopho-AKT (p-AKT) in HepG2-implated tissue were measured by immunohistochemistry. Tumor area, area of expression and expression ratio of pERK1/2 and p-AKT were determined.

Results: Increases in p-ERK1/2 and p-AKT expression in HepG2 group was related to changes in tumor growth in control, CUR, and THC groups. THC-treatment could attenuate the p-ERK1/2, p-AKT expression, tumor area, and ratio of expression in HepG2-implanted nude mice significantly, compared to CUR-treatment.

Conclusion: HepG2-induced tumor progression may be inhibited by THC in part through the inhibition of mitogen-activated protein kinase (MEK)/ERK and phosphoinositide 3-kinase (PI3K)/AKT.

Background: Vector control is a key strategy for eradication of filariasis, but it is limited, possibly due to rapid propagation from global warming. In Thailand, Mansonia mosquitoes are major vectors of filariasis caused by Brugia malayi filarial nematodes. However, little is yet known about vector biology and host-parasite relationship.

Objectives: Demonstrate the preliminary data of salivary gland morphology and protein profile of human filarial mosquitoes M. uniformis.

Methods: Morphology of M. uniformis salivary gland in both sexes was comparatively studied under a light microscope. Total protein quantization and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed to compare protein profile between male and female. In addition, quantitative analysis prior to and after blood feeding was made at different times (0, 12, 24, 36, 48, 60, and 72 hours).

Results: Total salivary gland protein of males and females was 0.32±0.03 and 1.38±0.02 μg/pair gland, respectively. SDS-PAGE analysis of the female salivary gland protein prior to blood meal demonstrated twelve bands of major proteins at 21, 22, 24, 26, 37, 39, 44, 53, 55, 61, 72, and 100 kDa. Compared to female, male salivary gland was composed of seven major protein bands at 39, 44, 53, 55, 61, 83, and 100 kDa. Quantitative study after blood feeding revealed that protein of 37 kDa decreased gradually whereas proteins of 61 and 83 kDa started to increase dramatically at 24 hours. It was postulated that the 37 kDa band, found only in the female, might serve as a candidate molecule for facilitating blood feeding.

Conclusion: Morphology and protein components of M. uniformis salivary glands might relate to blood feeding process and filarial disease transmission.

Background: Tinospora crispa (TC) has been used for years as diabetic remedy in Thai-traditional medicine. It has recently been reported that the aqueous extract from TC’s dried stem enhances glucose transport of L6 myotubes.

Objective: Investigate the effects of the water-ethanol extract of T. crispa (WETC) on glucose transport in a rat muscle cell-line L6 myotubes, and explore the mechanism of action of T. crispa in muscle cells.

Materials and methods: Glucose uptake rate was determined with the [³H] 2-deoxyglucose uptake method and the cell viability was assessed by sulforhodamine B (SRB) assay. Expressions of glucose transporter 1 (GLUT1), GLUT4, extracellular signal regulated kinase 1/2 (ERK1/2), and AMP-activated protein kinase (AMPK) were measured by quantitative Reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analysis.

Results: At 400 μg/mL, WETC significantly stimulated glucose uptake into L6 myotubes after 24 hours incubation (246.1 0.1% above basal; p <0.05). The enhancement of glucose transport by WETC was blocked by cycloheximide, a protein synthesis inhibitor. The up-regulation of GLUT1 protein was accompanied by a significant increased in GLUT1 mRNA expression. ERK1/2 was significantly activated in WETC-treated L6 myotubes. In addition, WETC treatment increased AMPK activity in L6 myotubes.

Conclusion: T. crispa increased glucose transport activity of L6 myotubes by enhancing GLUT1 expression, a result of AMPK stimulation.

Background: Russell’s viper venom-factor X activator (RVV-X) is a major procoagulant in Russell’s viper venom, and is composed of a heavy chain (RVV-XH) and two light chains (RVV-XL). It directly activates factor X in the final common coagulation pathway, which leads to rapid formation of blood clots.

Objective: Produce rabbit anti-recombinant protein antibodies and identify their cross-reactivity with two viperine snake venoms.

Methods: cDNA clones encoding RVV-XH and one of the light chains (RVV-XL; LC1) were recombinantly expressed in E. coli BL21 and used as antigens for rabbit immunization. The cross-reactivity of these anti-recombinant protein antibodies with two viperine snake venoms was determined using Western blot analysis.

Results: rRVV-XH was more immunogenic than rRVV-XL. Rabbit anti-rRVV-XH and rRVV-XL IgG antibodies bind specifically to RVV-X, but they do not neutralize purified RVV-X. In addition, rabbit anti-rRVV-XH IgG antibody also bind to an 18-kDa protein in C. rhodostoma venom, and many proteins in C. albolabris venom. Rabbit antirRVV- XL IgG antibody recognized protein bands of crude venoms of C. rhodostoma and C. albolabris at about 25-kDa and 23-kDa, respectively.

Conclusion: Rabbit anti-rRVV-XH and rRVV-XL IgG antibodies cross-reacted with molecules in other viperine venoms, which could have molecules with similar antigenic determinants. These antibodies could be useful to purify snake venom molecules by affinity chromatography as the first step in purification of factor X activator and other cross reacting molecules.

Background: Viral vectors can produce longer-lasting effects than a recombinant protein.

Objective: Develop a system of sustained GH1 (a human growth hormone gene) transfer and expression in baby hamster kidney cell (BHK-21) line using recombinant adeno-associated viral vectors pseudotyped with viral capsids from serotype 1 (rAAV2/1).

Methods: The expression of GH1 in vitro was examined by reverse transcription polymerase chain reaction and Western blot analysis. Effects of GH1 on cell proliferation were measured by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl-tetrazolium bromide (MTT) experiments. Mice serum IGF-1 and blood glucose, after injection of virus infected BHK-21 cells, were measured by enzyme-linked immunosorbent assays.

Results: rAAV2/1-mediated GH1 gene transfer occurred effectively in vitro after 48 hours of transduction at 1 10⁵ vg/mL (multiplicities of infection). MTT experiments indicated notable effects of the GH1 gene on cell proliferation in vitro. Subsequent animal experiment suggested that the injection of virus infected BHK-21 cells might induce increase of serum IGF-1.

Conclusion: Our study shows the feasibility of rAAV2/1-mediated GH gene delivery in vitro, applicable for future experimental and clinical investigations.

Background: Hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections are major health problems associated with blood transfusion practices in Pakistan. This study was conducted on a large population to asses the epidemiology of HCV and HIV in the North West Frontier Province (N.W.F.P) of Pakistan.

Methods: Between January 2008 and July 2009, 62,251 healthy blood donors (BDs), aged 17-50, were included. In that group, 61,059 (98.1%) were male and 1192 (1.9%) were female. All donors from the 11 areas of N.W.F.P were screened for HCV and HIV antibodies by ELISA (Biokit). Blood groups were also determined.

Results: HCV prevalence was 2.6% while HIV infection was 0.045% in this Pakistani population. Prevalence of HCV and HIV in 2008 was 2.4% and 0.049% respectively. In 2009, it was 3.0% for HCV and 0.038 for HIV. Among the age groups, the prevalence of HCV and HIV was not significantly different (p=0.128). Only 1484 (2.4%) of the donors were voluntary, the remaining were paid and family blood donors. The difference in the number of male and female donors was highly significant (p=0.00001). The B+ blood group was found in 30.5% followed by O+ in 25.9%, A+ in 24.9%, AB+ in 10.4%, A- in 2.6%, B- in 2.5%, O- in 2.3%, and AB- in 0.9%. Our study revealed a higher prevalence of HIV than in most of previous reports.

Conclusion: The frequency of HCV infection in blood donors is higher in N.W.F.P than in most of the rest of the world and lower than in other regions of Pakistan. Transfusion of infected blood is a common cause of transmission. HIV prevalence is increasing in Pakistan.

Background: Plateletpheresis procedures are thought to be generally safe without serious complications to the donor, and the products obtained are preferred over platelets derived from whole blood. However, safety issues about the donors have not been fully explored.

Objective: Investigate changes in hematological values after plateletpheresis in healthy volunteer donors.

Methods: A retrospective study was performed in 76 healthy donors at the Transfusion Medicine Unit, Universiti Sains Malaysia between 2004 and 2009. Haemonetics MCS+ and Trima accel (Gambro BCT) separators were used during plateletpheresis. Pre- and post-donation hematological values such as Hb, Hct, platelet, and leukocyte counts were measured using Sysmex XE-2100.

Results: After each procedure, there was a significant reduction in hemoglobin (pre-donation: 14.9g/dl, postdonation: 14.7g/dL), hematocrit (pre-donation: 44.6%, post-donation: 44.1%), platelet count (pre-donation: 264.0x10⁹/L, post-donation: 193.4x10⁹/L), mean platelet volume (pre-donation: 10.0fL, post-donation: 9.7fL), and platelet distribution width (pre-donation: 12.3fL, post-donation: 11.8fL).

Conclusion: There were significant drops in the donors hematological values post plateletpheresis.

Background: Lateral internal sphincterotomy has been accepted as the gold standard for chronic anal fissure. Despite the effectiveness, it carries a risk of fecal incontinence. Non-operative management of chronic anal fissure has been used as alternative treatment.

Objectives: Evaluate the efficacy and long-term outcomes of botulinum toxin injection for the treatment of chronic anal fissure.

Material and method: A prospective randomized controlled trial was conducted on 40 patients diagnosed with chronic anal fissure who were assigned to undergo either botulinum toxin injection or lateral internal sphincterotomy.

Results: The wound-healing rate at 12 weeks and the reduction of post-treatment-resting pressure was significantly better in the sphincterotomy group. Complications and the continence status were not different between the two treatment groups. None of the patients in the sphincterotomy group required additional treatment, while seven patients in the botulinum toxin injection group underwent subsequent sphincterotomy within three years.

Conclusion: The long-term result of botulinum toxin injection for chronic anal fissure was inferior to lateral internal sphincterotomy.

Background: Liver size can be physically evaluated by means of percussion, but measurement remains only gross-estimation. Ultrasonography is a non-invasive and accurate method for measurement of liver size. However, there are few ultrasonic data available for assessment of liver span in Thai children under two years old.

Objective: Determine liver sizes in healthy Thai children aged zero to two years and compare physical and ultrasonographic estimations.

Subject and method: Two hundred eighty one children (148 boys and 133 girls) were enrolled in this study. The age, weight, and height were assessed, and the body surface area was calculated. The liver size or liver span was estimated by percussion and compared to ultrasonography.

Result: The liver span (mean±SD) was 5.4±1.0 cm, 5.1±1.1 cm by physical and ultrasonographic examination, respectively. The measurement of liver span by physical examination had high reliability. The liver span correlated with body surface area rather than height, age, and weight.

Conclusion: Physical examination of liver span provided reliable data in Thai children under two years old.

Background: Mild traumatic brain injury (TBI) is a common neurological disorder. Cognitive impairment is a result of TBI, and executive function is impaired in various degree of injury. Few data are available for assessment of executive dysfunction in Thai patients.

Objective: Examine prevalence and factors influencing executive dysfunction among mild TBI patients in Thailand.

Materials and methods: A cross-sectional descriptive study was conducted to determine the prevalence of 60 mild TBI patients. The patients were selected from those who were admitted at Khon Kaen Hospital between September and December 2009. Patients with previously major psychiatric or neurological disorders and currently confused or depressed were excluded. The executive function was measured by the Wisconsin Card Sorting Test (WCST), and the data was analyzed statistically.

Results: Prevalence of executive dysfunction among the patients was 21.7%. Primary education level and low monthly salary were two factors to be associated with dysfunction. Alcohol use was associated with WCST score, but not with the dysfunction. However, severity of injury had no significant association with both the dysfunction and WCST score.

Conclusion: Executive dysfunction in mild TBI patients was high (21.7% prevalent in Northeastern Thailand). Primary education level and low monthly salary were associated with dysfunction.

Background: Carboxymethylchitosan is a chitosan-derivative obtained from the carboxymethylation of chitin with chloroacetic acid in alkaline solution. It has shown its potential in animal model studies as an accelerator of wound healing.

Materials and methods: Prospective, comparative clinical trials of traditional tulle gauze, alginate membrane, and carboxymethylchitosan sponge were carried out in the treatment of partial- thickness skin graft donor sites. Between June 2005 and March 2006, 70 donor sites from 44 patients were randomly treated by these three different wound dressing materials. Each wound was treated until it was completely healed, and a visual analogue scale was used for the pain evaluation.

Results: The results showed that the donor sites dressed with carboxymethylchitosan or alginate healed more rapidly than those treated with tulle gauze. There was no significant difference in the healing rate between carboxymethylchitosan and alginate. The pain scores evaluated among these three dressing groups did not significantly differ.

Conclusion: Carboxymethylchitosan is as efficacious as traditional tulle gauze or alginate membrain in the treatment of partial thickness skin graft donor sites.

Background: Klinefelter syndrome may present as precocious puberty, which can be either central precocious puberty or peripheral precocious puberty, caused by an extragonadal germ cell tumor.

Objective: Report two cases of Klinefelter syndrome that presented with precocious puberty due to a β-hCG producing mediastinal tumor.

Method: Review of the clinical history, physical examination, and laboratory investigations.

Results: Pseudo-precocity developed some years before diagnosis of -hCG producing tumor. The patients did not have typical physical features of this syndrome. The testes were small and had loose consistency.

Conclusion: Klinefelter syndrome must be excluded in all boys presenting with precocious puberty due to a β-hCG producing tumor. Conversely, patients with Klinefelter syndrome should be regularly checked for β-hCG and α-fetoprotein levels. In those cases, the patients can be diagnosed and treated early. With the early treatment, they will be able to attain normal adult height and have fewer complications from the tumor.

Background: There has been no report of dog rabies in Sultanate of Oman, possibly due to an inadequate active and passive surveillance and response system.

Methods: We recently developed Fluorescent Antibody Test (FAT) and real time polymerase chain reaction (PCR) as a sensitive diagnosis of rabies.

Results: We present the first laboratory proven canine rabies case in Sultanate of Oman. Laboratory facilities for sensitive diagnosis are now available in Sultanate of Oman.

Conclusion: A systematic surveillance system for rabies in domesticated dogs will have a good influence on the control of this zoonotic infection in Sultanate of Oman.