Chairat Tantrawatpan

Comparison of endothelial progenitor cell function in type 2 diabetes with good and poor glycemic control

BackgroundEndothelial progenitor cells (EPCs) play an important role in vascular repair and a decrease in the number of EPCs is observed in type 2 diabetes. However, there is no report on the change of EPCs after glycemic control. This study therefore aimed to investigate the EPC number and function in patients with good and poor glycemic control.MethodsThe number of EPCs was studied using flow cytometry by co-expression of CD34 and VEGFR2. The EPCs were cultured and characterized by the expression of UEA-I, CD34, VEGFR2, vWF and Dil-Ac-LDL engulfment, as well as the ability to form capillary-like structures. An in vitro study on the effect of hyperglycemia on the proliferation and viability of the cultured EPCs was also performed.ResultsThe number of EPCs in type 2 diabetes was significantly decreased compared with healthy controls and there was an inverse correlation between the EPC numbers and plasma glucose, as well as HbA1C. The number and function of EPCs in patients with good glycemic control were recovered compared with those with poor glycemic control. When glucose was supplemented in the culture in vitro, there was a negative effect on the proliferation and viability of EPCs, in a dose-dependent manner, whereas the enhancement of apoptosis was observed.ConclusionThere was EPC dysfunction in type 2 diabetes which might be improved by strict glycemic control. However, the circulating EPC number and proliferative function in patients with good glycemic control did not reach the level in healthy controls.

Immunosuppressive properties of mesenchymal stromal cells derived from amnion, placenta, Wharton's jelly and umbilical cord

The role of bone marrow‐derived mesenchymal stromal cells (BM‐MSC) in preventing the incidence and ameliorating the severity of graft‐versus‐host disease (GvHD) has recently been reported. However, as the collection of BM‐MSC is an invasive procedure, more accessible sources of MSC are desirable.

Evaluation of Immunoglobulin G Subclass Antibodies against Recombinant Fasciola gigantica Cathepsin L1 in an Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Human Fasciolosis

ABSTRACT A cystatin capture enzyme-linked immunosorbent assay (ELISA) using recombinant Fasciola gigantica cathepsin L1 antigen was developed to detect specific immunoglobulin G (IgG) subclass antibodies (IgG1, IgG2, IgG3, and IgG4) and was evaluated for its diagnostic potential for human fasciolosis. In an analysis of the sera of 13 patients infected with F. gigantica, 209 patients with other parasitic infections, 32 cholangiocarcinoma patients, and 42 healthy controls, the IgG4-ELISA gave the highest diagnostic values. The sensitivity, specificity, accuracy, and positive and negative predictive values of this method based on the detection of IgG4 antibody were 100%, 99.3%, 99.3%, 86.7%, and 100%, respectively. The results revealed that restricting the ELISA to the detection of specific IgG4 antibody enhanced the specificity and accuracy for the serodiagnosis of human fasciolosis.

Molecular identification of Paragonimus species by DNA pyrosequencing technology

DNA pyrosequencing for PCR amplicons is an attractive strategy for the identification of microorganisms because of its short time performance for large number of samples. In this study, the primers targeting the fragment of ITS2 region of nuclear ribosomal RNA gene were newly developed for pyrosequencing-based identification of 6 Paragonimus species, Paragonimus bangkokensis, Paragonimus harinasutai, Paragonimus heterotremus, Paragonimus macrorchis, Paragonimus siamensis and Paragonimus westermani. Pyrosequencing determination of 39 nucleotides of partial ITS2 region could discriminate 6 Paragonimus species, and could also detect intra-species genetic variation of P. macrorchis. This DNA pyrosequencing-based identification can be a valuable tool to improve species-level identification of Paragonimus in the endemic areas.

Zoonotic Echinostome Infections in Free-Grazing Ducks in Thailand

Free-grazing ducks play a major role in the rural economy of Eastern Asia in the form of egg and meat production. In Thailand, the geographical location, tropical climate conditions and wetland areas of the country are suitable for their husbandry. These environmental factors also favor growth, multiplication, development, survival, and spread of duck parasites. In this study, a total of 90 free-grazing ducks from northern, central, and northeastern regions of Thailand were examined for intestinal helminth parasites, with special emphasis on zoonotic echinostomes. Of these, 51 (56.7%) were infected by one or more species of zoonotic echinostomes, Echinostoma revolutum, Echinoparyphium recurvatum, and Hypoderaeum conoideum. Echinostomes found were identified using morphological criteria when possible. ITS2 sequences were used to identify juvenile and incomplete worms. The prevalence of infection was relatively high in each region, namely, north, central, and northeast region was 63.2%, 54.5%, and 55.3%, respectively. The intensity of infection ranged up to 49 worms/infected duck. Free-grazing ducks clearly play an important role in the life cycle maintenance, spread, and transmission of these medically important echinostomes in Thailand.

Molecular Markers for Detection and Differentiation of Plasmodium falciparum and Plasmodium vivax in Human Blood Samples by Pyrosequencing

ABSTRACT PCR amplification coupled with pyrosequencing was used to measure molecular markers that could be used to detect and differentiate Plasmodium falciparum and Plasmodium vivax in human blood samples. The detection rates were in agreement with the results of Giemsa-stained film microscopy, which is the current gold standard for detection. This method provides an exciting alternative for malaria diagnosis.

Evaluation of immunoglobulin G4 subclass antibody in a peptide-based enzyme-linked immunosorbent assay for the serodiagnosis of human fascioliasis.

SUMMARYTo improve the diagnosis of human fascioliasis caused by Fasciola gigantica, we developed a peptide-based enzyme-linked immunosorbent assay (peptide-based ELISA) based on the detection of specific IgG4 subclass antibody. Two identified B-cell epitopes of F. gigantica cathepsin L1 were synthesized as single synthetic peptides, acetyl-DKIDWRESGYVTELKDQGNC-carboxamide (peptide L) and acetyl-DKIDWRESGYVTEVKDQGNC-carboxamide (peptide V), and their diagnostic potential was evaluated. The sera of 25 patients infected with F. gigantica, 212 patients with other parasitic infections, 32 cholangiocarcinoma patients and 57 healthy controls were analysed. The sensitivity, specificity, accuracy, and positive and negative predictive values of this assay were the same with both peptides at 100%, 99.7%, 99.7%, 96.2% and 100%, respectively. These highly sensitive and specific peptide-based ELISAs for the detection of specific IgG4 antibody could be useful for laboratory diagnosis of human fascioliasis in future large-scale surveys throughout Southeast Asia where this disease is prevalent.

Molecular Identification of Trichinella papuae from a Thai Patient with Imported Trichinellosis

Previously, we reported the presence of imported trichinellosis in a Thai worker returning from Malaysia, who presented with progressive generalized muscle hypertrophy and weakness after eating wild boar meat. This work analyzed a partial small subunit of a mitochondrial ribosomal RNA gene of Trichinella larvae isolated from the patient. The results showed complete identity with a mitochondrial RNA gene of Trichinella papuae (GenBank accession no. EF517130). This is the first report of imported trichinellosis in Thailand caused by T. papuae. It is possible that T. papuae is widely distributed in the wildlife of Southeast Asia.

Genetic characterization of Echinostoma revolutum and Echinoparyphium recurvatum (Trematoda: Echinostomatidae) in Thailand and phylogenetic relationships with other isolates inferred by ITS1 sequence

Echinostomatidae are common, widely distributed intestinal parasites causing significant disease in both animals and humans worldwide. In spite of their importance, the taxonomy of these echinostomes is still controversial. The taxonomic status of two species, Echinostoma revolutum and Echinoparyphium recurvatum, which commonly infect poultry and other birds, as well as human, is problematical. Previous phylogenetic analyses of Southeast Asian strains indicate that these species cluster as sister taxa. In the present study, the first internal transcribed spacer (ITS1) sequence was used for genetic characterization and to examine the phylogenetic relationships between an isolate from Thailand with other isolates available from GenBank database. Interspecies differences in ITS1 sequence between E. revolutum and E. recurvatum were detected at 6 (3%) of the 203 alignment positions. Of these, nucleotide deletion at positions 25, 26, and 27, pyrimidine transition at 50, 189, and pyrimidine transversion at 118 were observed. Phylogenetic analysis revealed that E. recurvatum from Thailand clustered as a sister taxa with E. revolutum and not with other members of the genus Echinoparyphium. Interestingly, this result confirms a previous report based on allozyme electrophoresis and mitochondrial DNA that E. revolutum and E. recurvatum in Southeast Asia are sister species. Hence, the taxonomic status of E. recurvatum in Thailand, as well as in Southeast Asian countries needs to be confirmed and revised using more comprehensive analyses based on morphology and other molecular techniques.

Spatial and temporal genetic variation of Echinostoma revolutum (Trematoda: Echinostomatidae) from Thailand and the Lao PDR

A total of 314 individual Echinostoma revolutum were collected at different locations and times from domestic ducks from Khon Kaen Province, Thailand and Vientiane Province, the Lao Peoples Democratic Republic (PDR). Genetic variation of these parasites was analyzed using multilocus enzyme electrophoresis at three polymorphic loci namely, glucose-6-phosphate dehydrogenase (G6pd), malic enzyme (Me) and peptidase valine-leucine (PepA). High levels of genetic variability were found within and between populations. Significant heterozygote deficiencies compared with the predictions under Hardy-Weinberg equilibrium were detected in populations from Thailand and the Lao PDR for all loci except G6pd-1. Significant genetic differentiation was observed between spatially separated populations from Thailand and the Lao PDR. This as also true for some samples collected at different times in Thailand. The variability found may be consistent with a Wahlund effect, genetic drift and/or other factors such as the population structure of snail hosts. Our data provide further insight into the process of genetic divergence within and among geographically and temporally isolated populations of E. revolutum, and potentially other medically important echinostomes in Southeast Asia.