Tanawan Kummalue

Cytotoxic properties of root extract and fruit juice of Trichosanthes cucumerina.

The root extract of Trichosanthes cucumerina L. and bryonolic acid (1), its main constituent, as well as the fruit juice and cucurbitacin B (3), its main constituent, were tested for cytotoxicity against four human breast cancer cell lines (SKBR3, MCF7, T47D, and MDA-MB435), two lung cancer cell lines (A549 and SK-LU1), and one colon cancer cell line (Caco-2). The root extract had higher IC (50) values than bryonolic acid (1) against three breast cancer cell lines (MCF7 = 267/121, T47D = 316/124, MDA-MB435 = 140/90 microL/mL) and one lung cancer cell line (A549 = 106/100 microL/mL). The fruit juice also had higher IC (50) values than cucurbitacin B (3) against the four breast cancer cell lines (131/73, 375/35, 249/60, and 156/26 microL/mL, respectively) and one lung cancer cell line (141/41 microL/mL) as shown above, as well as against the colon cancer cell line (101/1.5 microL/mL). However, the root extract inhibited SK-LU1 more strongly than did the fruit juice, cucurbitacin B (3), and bryonolic acid (1) (149/169/180/>500 microL/mL, respectively). The root extract inhibited the two lung and three breast cancer cell lines (SKBR3, MDA-MB435, and MCF7) more strongly than the fruit juice. Bryonolic acid (1) inhibited MDA-MB435 somewhat better than the other tested human cancer cell lines. The fruit juice inhibited the colon cancer cell line (Caco-2) more strongly than the root extract. Cucurbitacin (3) inhibited human cancer cell lines, especially Caco-2, much more strongly than bryonolic acid (1). In addition to bryonolic acid (1), bryononic acid (2), cucurbitacin B (3), and dihydrocucurbitacin B (4) also were isolated from the root extract.

Aberrant antigenic expression in extranodal NK/T-cell lymphoma: a multi-parameter study from Thailand

BackgroundExtranodal NK/T-cell lymphoma, nasal type (ENKTL) is not common worldwide, but it is the most common T- and NK-cell lymphomas in many Asian countries. Immunophenotypic profiles were studied based on limited series. The authors, therefore, studied on ENKTL according to characterize immunophenotypic profiles as well as the distribution of EBV subtype and LMP-1 gene deletion.MethodsBy using tissue microarray (TMA), immunohistochemical study and EBV encoded RNA (EBER) in situ hybridization were performed. T-cell receptor (TCR) gene rearrangement, EBV subtyping, and LMP-1 gene deletion were studied on the available cases.ResultsThere were 22 cases eligible for TMA. ENKTL were positive for CD3 (91%), CD5 (9%), CD7 (32%), CD4 (14%), CD56 (82%), TIA-1 (100%), granzyme B (95%), perforin (86%), CD45 (83%), CD30 (75%), Oct2 (25%), and IRF4/MUM1 (33%). None of them was positive for βF1, CD8, or CD57. TCR gene rearrangement was negative in all 18 tested cases. EBV was subtype A in all 15 tested cases, with 87% deleted LMP-1 gene. Cases lacking perforin expression demonstrated a significantly poorer survival outcome (p = 0.008).ConclusionsThe present study demonstrated TIA-1 and EBER as the two most sensitive markers. There were a few CD3 and/or CD56 negative cases noted. Interestingly, losses of CD45 and/or CD7 were not uncommon while Oct2 and IRF4/MUM1 could be positive in a subset of cases. Based on the present study in conjunction with the literature review, determination of PCR-based TCR gene rearrangement analysis might not be a useful technique for making diagnosis of ENKTL.

Detection of monoclonal immunoglobulin heavy chain gene rearrangement (FR3) in Thai malignant lymphoma by High Resolution Melting curve analysis

AbstractMalignant lymphoma, especially non-Hodgkin lymphoma, is one of the most common hematologic malignancies in Thailand. The diagnosis of malignant lymphoma is often problematic, especially in early stages of the disease. Detection of antigen receptor gene rearrangement including T cell receptor (TCR) and immunoglobulin heavy chain (IgH) by polymerase chain reaction followed by heteroduplex has currently become standard whereas fluorescent fragment analysis (GeneScan) has been used for confirmation test. In this study, three techniques had been compared: thermocycler polymerase chain reaction (PCR) followed by heteroduplex and polyacrylamide gel electrophoresis, GeneScan analysis, and real time PCR with High Resolution Melting curve analysis (HRM). The comparison was carried out with DNA extracted from paraffin embedded tissues diagnosed as B- cell non-Hodgkin lymphoma. Specific PCR primers sequences for IgH gene variable region 3, including fluorescence labeled IgH primers were used and results were compared with HRM. In conclusion, the detection IgH gene rearrangement by HRM in the LightCycler System showed potential for distinguishing monoclonality from polyclonality in B-cell non-Hodgkin lymphoma.IntroductionMalignant lymphoma, especially non-Hodgkin lymphoma, is one of the most common hematologic malignancies in Thailand. The incidence rate as reported by Ministry of Public Health is 3.1 per 100,000 population in female whereas the rate in male is 4.5 per 100,000 population [1]. At Siriraj Hospital, the new cases diagnosed as malignant lymphoma were 214.6 cases/year [2]. The diagnosis of malignant lymphoma is often problematic, especially in early stages of the disease. Therefore, detection of antigen receptor gene rearrangement including T cell receptor (TCR) and immunoglobulin heavy chain (IgH) by polymerase chain reaction (PCR) assay has recently become a standard laboratory test for discrimination of reactive from malignant clonal lymphoproliferation [3, 4]. Analyzing DNA extracted from formalin-fixed, paraffin-embedded tissues by multiplex PCR techniques is more rapid, accurate and highly sensitive. Measuring the size of the amplicon from PCR analysis could be used to diagnose malignant lymphoma with monoclonal pattern showing specific and distinct bands detected on acrylamide gel electrophoresis. However, this technique has some limitations and some patients might require a further confirmation test such as GeneScan or fragment analysis [5, 6].GeneScan technique or fragment analysis reflects size and peak of DNA by using capillary gel electrophoresis. This technique is highly sensitive and can detect 0.5-1% of clonal lymphoid cells. It measures the amplicons by using various fluorescently labeled primers at forward or reverse sides and a specific size standard. Using a Genetic Analyzer machine and GeneMapper software (Applied Bioscience, USA), the monoclonal pattern revealed one single, sharp and high peak at the specific size corresponding to acrylamide gel pattern, whereas the polyclonal pattern showed multiple and small peak condensed at the same size standard. This technique is the most sensitive and accurate technique; however, it usually requires high technical experience and is also of high cost [7]. Therefore, rapid and more cost effective technique are being sought.LightCycler PCR performs the diagnostic detection of amplicon via melting curve analysis within 2 hours with the use of a specific dye [8, 9]. This dye consists of two types: one known as SYBR-Green I which is non specific and the other named as High Resolution Melting analysis (HRM) which is highly sensitive, more accurate and stable. Several reports demonstrated that this new instrument combined with DNA intercalating dyes can be used to discriminate sequence changes in PCR amplicon without manual handling of PCR product [10, 11]. Therefore, current investigations using melting curve analysis are being developed [12, 13].In this study, three different techniques were compared to evaluate the suitability of LightCycler PCR with HRM as the clonal diagnostic tool for IgH gene rearrangement in B-cell non-Hogdkin lymphoma, i.e. thermocycler PCR followed by heteroduplex analysis and PAGE, GeneScan analysis and LightCycler PCR with HRM.

Significant Increase in Cytotoxic T Lymphocytes and Natural Killer Cells by Triphala: A Clinical Phase I Study

Background. Searching for drugs or herbal formulations to improve the immunity of HIV/AIDS positive people is an important issue for researchers in this field. Triphala, a Thai herbal formulation, is reported to have immunomodulatory effects in mice. However, it has not yet been investigated for immunostimulatory and side effects in healthy human volunteers. Objective. To evaluate the immunostimulatory and side effects of Triphala in a clinical phase I study. Materials and Methods. All volunteers took Triphala, 3 capsules per day for 2 weeks. Complete physical examination, routine laboratory analysis, and immunological studies were performed before ingestion and after initial meeting for 4 consecutive weeks. Results. We found that Triphala demonstrated significant immunostimulatory effects on cytotoxic T cells (CD3−CD8+) and natural killer cells (CD16+CD56+). Both of them increased significantly when compared with those of the control samples. However, no significant change in cytokine secretion was detected. All volunteers were healthy and showed no adverse effects throughout the duration of the study. Conclusion. Triphala has significant immunostimulatory effects on cellular immune response, especially cytotoxic T cells and natural killer cells. Increases in the absolute number of these cells may provide a novel adjuvant therapy for HIV/AIDS positive people in terms of immunological improvement.

Antibacterial and Antiproliferative Activities of Plumericin, an Iridoid Isolated from Momordica charantia Vine

Plumericin, an iridoid lactone, was isolated with relatively high yield from Momordica charantia vine using the supercritical fluid extraction (SFE) and the separation box (Sepbox) comprising dual combination of high-performance liquid chromatography and solid phase extraction. This compound showed antibacterial activity against Enterococcus faecalis and Bacillus subtilis with minimum inhibitory concentration (MIC) values better than cloxacillin. Plumericin potently inhibited proliferation of two leukemic cancer cell lines: they were acute and chronic leukemic cancer cell lines, NB4 and K562, with the effective doses (ED50) of 4.35 ± 0.21 and 5.58 ± 0.35 μg/mL, respectively. In addition, the mechanism of growth inhibition in both cell lines was induced by apoptosis, together with G2/M arrest in K562 cells.

Antiproliferative Effect and the Isolated Compounds of Pouzolzia indica.

Previous report showed the high potent antiproliferative effect of the methanolic part extracted from the aerial parts of Pouzolzia indica on NB4 and HT93A acute leukemic cell lines with the IC50 values of 28.5 and 49.8 μg/mL, respectively. The bioassay-guided fractionation of the methanolic part gave 5 fractions, that is, FFI–FFV. FFII, FFIII, and FFIV inhibited the above leukemic cell lines with the IC50 values of 15.1 (FFII), 14.4 (FFIII), 32.1 (FFIV), and 31.0 (FFII), 9.7 (FFIII), 10.5 (FFIV) μg/mL, respectively. The compounds in these fractions were isolated using chromatographic technique. FFII contained friedelin 1, 28-hydroxy-3-friedelanone 2, and 7-methoxy-coumarin 3. FFIII contained 6, 7-dimethoxy-coumarin 4, scopoletin 5, methyl caffeate 6. FFIV contained sitosteryl glucoside 7 and a supposed glycosphingolipid 8. The chemical structures were elucidated by spectroscopic methods.

Cutaneous involvement by colonic extranodal NK/T-cell lymphoma mimicking mycosis fungoides: a case report*.

We report a 51‐year‐old woman with cutaneous involvement by extranodal NK/T‐cell lymphoma (TCL) of the colon that microscopically mimicked mycosis fungoides (MF). She had a history of fever of unknown origin for 2 months and then developed multiple erythematous papules on her trunk and extremities. A skin biopsy revealed superficial infiltration by atypical small to medium‐sized lymphocytes with epidermotropism and Pautrier collections. Immunohistochemical studies showed expression of CD3 and TIA‐1 with lack of expression (double negative) of CD4 and CD8. Initially, we reported the diagnosis as MF, cytotoxic variant. Thereafter, computerized tomography scan incidentally identified a colonic mass. A colonic biopsy revealed infiltration of atypical lymphoid cells with the same morphology and immunophenotype as those found in the skin. Additionally, CD56 and Epstein‐Barr virus‐encoded RNA in situ hybridization in both skin and colonic biopsies were diffusely positive. Thus, extranodal NK/TCL was diagnosed. Delta T‐cell receptor (TCR) gene rearrangement was documented in the skin biopsy by polyacrylamide gel electrophoresis and fluorescence capillary gel electrophoresis methods. There was no TCR gene rearrangement detected in the colonic biopsy. Unfortunately, the patient died within 2 months of diagnosis.

CK19 Detection by Multiplex Real-Time RT-PCR Correlates with the Presenceof Angiolymphatic and Perineural Invasion in Sentinel Lymph Nodes of ThaiBreast Cancer Patients

Abstract Background: Pathological evaluation of sentinel lymph nodes of breast cancer patients is a critical factor in determining the treatment strategy including patients’ outcome. Multiple factors have been known to correlate with disease free survival and clinical outcomes such as sentinel lymph node metastases. Recently, real-time RT-PCR which is a very useful, rapid, high sensitivity and specificity tool has been reported to detect micrometastases in sentinel lymph nodes which could be helpful to identify high risk patients. Objectives: The purpose of this study are: To evaluate the sensitivity, specificity, and accuracy of CK19 detection by multiplex real-time RT-PCR and to elucidate the correlation of multiplex real-time RT-PCR results and clinicopathological parameters in sentinel lymph nodes of Thai breast cancer patients. Material and methods: 119 patients diagnosed as breast cancer stage I or II at Siriraj Hospital during October 2008 – June 2010 were enrolled in this study. RNAs were extracted from their sentinel lymph nodes and cDNAs were synthesized. Multiplex real-time RT-PCR for detection of CK19 was performed and analysed. Patients’ characteristics and pathological parameters were compared with multiplex real-time RT-PCR results. Results: Significant correlation between angiolymphatic and perineural invasion status and CK19 expression was demonstrated. The overall concordance of CK19 gene with pathological results was 72.3%. Sensitivity and specificity for CK19 detected by multiplex real-time RT-PCR were 78.3%, and 66.1%, respectively. Conclusion: Significant correlation between angiolymphatic and perineural invasion status and CK19 expression in sentinel lymph nodes of breast cancer patients was found in this study.

Ribosomal protein L11- and retinol dehydrogenase 11-induced erythroid proliferation without erythropoietin in UT-7/Epo erythroleukemic cells.

Erythropoiesis is the process of proliferation, differentiation, and maturation of erythroid cells. Understanding these steps will help to elucidate the basis of specific diseases associated with abnormal production of red blood cells. In this study, we continued our efforts to identify genes involved in erythroid proliferation. Lentivirally transduced UT-7/Epo erythroleukemic cells expressing ribosomal protein L11 (RPL11) or retinol dehydrogenase 11 (RDH11) could proliferate in the absence of erythropoietin, and their cell-cycle profiles revealed G0/G1 prolongation and low percentages of apoptosis. RPL11-expressing cells proliferated more rapidly than the RDH11-expressing cells. The antiapoptotic proteins BCL-XL and BCL-2 were expressed in both cell lines. Unlike the parental UT-7/Epo cells, the expression of hemoglobins (Hbs) in the transduced cells had switched from adult to fetal type. Several signal transduction pathways, including STAT5, were highly activated in transduced cells; furthermore, expression of the downstream target genes of STAT5, such as CCND1, was upregulated in the transduced cells. Taken together, the data indicate that RPL11 and RDH11 accelerate erythroid cell proliferation by upregulating the STAT5 signaling pathway with phosphorylation of Lyn and cyclic AMP response element-binding protein (CREB).

Botanicals in Dietary Supplements

Botanicals are accepted worldwide as medicinal agents and nutraceuticals. Extensive scientific investigations have been performed over the past 200 years which have resulted in the evolution of botanical utilization. This special issue highlights some of this research in 7 review articles and in 24 original research articles. Review Articles. W. Jiratchariyakul and G. B. Mahady wrote about the botanical status and popular herbs in EU, US, and Thailand. M. Miroddi et al. updated the market and the regulatory of botanical products in EU and US. Traditional Chinese and Indian Medicines (TCM and TIM) play an important role in Asian countries. Z. Wang et al. presented and discussed the role of TCM in the treatment of epidemic type II diabetes mellitus. M. M. Pandey et al. reported the use of TIM as a nutritional supplement in malnutrition. Y. Kamisah et al. presented the chemoprevention and antioxidation of Parkia speciosa. A. P. Bartolome et al. reviewed laboratory evidence of Bidens pilosa. T.-P. Huynh et al. discussed the promising botanical compounds for prevention and treatment of eye diseases. Research Articles. New biological and pharmacological activities of botanicals are reported. They included the enhancement of learning, memory and antistress of Acanthopanax trifoliatus (P. Sithisarn et al.), the sedative effect of Ziziphus mauritiana (A. M. M. San et al.), the cardioprotective effect of Phyllanthus emblica (L. Chularojmontri et al.), the cytoprotection and antioxidative stress of Citrus maxima (L. Chularojmontri et al.), anti-influenza viral activity of Momordica charantia (V. Pongthanapisith et al.), antioxidation from Nypa fruticans (N. Prasad et al.), Nigella glandulifera (J. Zhao et al.), TIM (M. M. Pandey et al.), Herba Cynomorii (J. Chen et al.). The alleviation of metabolic disorder of Citrus ichangensis (X. Ding et al.), and anticancer activity of Vitex agnus-castus (S. Li et al.). The mechanisms of anticancer action are deeply investigated with the botanical compounds, zerumbone from Zingiber zerumbet (N. M. Nadzri et al.), phenyl butenoid dimmer from Zingiber cassumunar (T. Anasamy et al.), and girinimbine from Murraya koenigii (S. Mohan et al.). Biochanin A, the major isoflavone from Trifolium pratense, prevented the bone loss in the ovariectomized rat (S.-J. Su et al.). Quercetin isolated from Caesalpinia mimosoides had neuroprotective effect (N. Tangsaengvit et al.) and inhibited eosinophile (M. K. Asano Sakai-Kashiwahara). C. Li et al. reported new anti-inflammatory triterpenoids from Illicium difengpi. Besides the herbal activities, the cohort study of TCM, Si-Wu-Tang, in postpartum women was performed to evaluate the health benefits. The effectiveness of the modern herbal drug was carried out using the double-blind randomized controlled clinical trial, as shown under the title “Antiherpetic effects of Gynura procumbens” (S. Jarikasem et al.). The research on the quality assessment of the botanicals was also presented in this issue. It included the HPLC analysis of Moringa oleifera (B. Vongsak et al.) and Pueraria tuberosa (S. Rastogi et al.). In addition, the arsenic accumulation in Zingiberaceous rhizomes was reported (C. Ubonnuch et al.). Weena Jiratchariyakul Ludger Beerhues Gail B. Mahady Tanawan Kummalue Molvibha Vongsakul