Recombinant Adenovirus Expressing Anti-Cancer Gene in Colon Cancer Cell Explant in Mice

Gene therapy is an alternative method to cure or slow down the progression of malignant cancer. Recombinant adenovirus encoding viral protein 2 (VP2) (ADV-VP2) of very virulent infectious bursal disease virus (vvIBDV) was employed to eliminate cancer cells by apoptosis mechanism. Besides, another recombinant adenovirus encoding murine endostatin (ADV-endo) was constructed aiming to block the formation of new blood vessels that supply nutrients to tumor. Recombinant adenoviruses were found to express the VP2 gene at a significantly high level in cancer cells, especially adenocarcinomas, with the relative quantification (RQ) value from 149.58 to 233.12 fold 72 hour (hr) post-infection (p.i). However, only small traces of VP2 gene expression was found in non-cancer cells, with the RQ value ranging from 0.04 to 0.54 fold 72 hr p.i. The capacity of recombinant adenovirus to infect target cells is dependent on the level of coxsackievirus adenovirus receptor (CAR) available in each cell. DNA fragmentation test, TUNEL assay, FITC Annexin V/PI double staining quantification test and caspase tests were carried out to determine the apoptosis induction level by recombinant adenovirus as well as the apoptosis related pathway. All four apoptosis tests were in agreement with recombinant adenovirus induced apoptosis in cancer cells, particularly in MCF-7, CT26 and HepG2, but not in non-cancer cells. CT26 cells demonstrated DNA fragmentation as early as 24 hr p.i, followed by MCF-7 and HepG2 cells, which showed DNA fragments during 48 and 72 hr p.i. These three cancer cells indicated significantly higher apoptotic cells proportion via TUNEL assay and FITC Annexin V/PI double staining test, with the percentage of apoptotic cells ranging from 78.0% to 60.0%. Caspase tests indicated that recombinant adenovirus activated apoptosis at the late stage of infection, through the intrinsic pathway by caspase 2 (initiator caspase), then led to the activation caspase 3 (effector caspase). No apoptosis was detected in cancer cells infected with mock adenovirus vector, thus apoptosis induction was solely contributed by the inserted gene. Colon cancer cells explanted mice were used as a model for cancer therapy in the present study. Tumor size regression was found in multiple doses of recombinant adenovirus treated mice but no regression was found in control mice. Partial tumor size regression was observed in mice treated with 1 dose of ADV-VP2. Complete regression of tumor mass was observed in 5 out of 6 mice and 2 out of 6 mice treated with 3 and 2 doses of ADV-VP2, respectively. Combined treatment of ADV-VP2 and ADV-endo demonstrated prolong mice survival time for up to one month as compared to control mice. Female mice can survive 15 days longer than male mice which suffered from similar large tumor mass. Mouse organs of recombinant adenovirus treated groups were comparable to the control group due to the nature of adenovirus which transiently expressed. The gene expression level in mouse intestines were significantly higher than other organs, 93.06±1.82 fold in 3 doses ADV-VP2 treated mice. Findings collectively justified the ability of ADV-VP2 to induce apoptosis effectively in tumor mass upon booster administration. In conclusion, the combined administration of recombinant adenovirus (ADV-VP2 and ADV-endo) had therapeutic potential against cancer. Further investigation on the optimal dosage of combined therapy need to be carried out in order to achieve the augmentative effect of these constructs on cancer therapy

Phytochemicals derived from soya bean husk exert hypoglycemic and anti-adipogenic properties in cell culture models

Purpose: Literature has shown that phenolic acids and flavonoids are bearing with hypoglycemic and anti-adipogenic properties. Therefore, this study aims to evaluate the possibility of phenolic-rich soya bean husk powder extract (SHPE) in combating diabetes and obesity using in vitro models. Design/methodology/approach: The hypoglycemic properties were evaluated by determining the ability of SHPE (25-100 µg/mL) in inhibiting a-amylase and a-glucosidase enzymes and in triggering insulin secretion in BRIN-BD11 cells. Murine 3T3-L1 adipocytes were used for evaluating the anti-adipogenic properties of SHPE through the determination of relative lipid accumulation, triglyceride content and glycerol-3-phosphate dehydrogenase (GPDH) activity. Findings: The hypoglycemic properties of SHPE was in the dose-dependent manner, where 100 µg SHPE/mL exhibited a significant higher (p < 0.05) a-amylase inhibitory activity (56.8 ± 0.11 per cent) and insulin secretion activity (0.73 ± 0.02 µg/l) against other concentrations. In contrast to the aforementioned findings, a significant lower a-glucosidase inhibitory activity (52.0 ± 0.44 per cent) was also observed in 100 µg SHPE/mL. Nevertheless, findings revealed that all the SHPE were able to inhibit the activity of a-amylase and a-glucosidase and stimulated the insulin secretion in BRIN-BD11 cells. On the other hand, the anti-adipogenic properties of SHPE were in the reverse dose-dependent manner, where 100 µg SHPE/mL demonstrated a significant lower (p < 0.05) relative lipid accumulation (48.5 ± 0.03 per cent), intracellular triglyceride content (5.7 ± 0.07 mg/dL) and GPDH activity (1.0 ± 0.01 mU/mL). These findings reflected that 100 µg SHPE/mL was a potent anti-adipogenic agent when compared with other concentrations. In conclusion, soya husk could emerge as a potential hypoglycemic and anti-adipogenic agents in in vitro models. Originality/value: This was the first study to explore the effectiveness of phytochemicals derived from soya bean husk in ameliorating hyperglycemia and adipogenesis. Promising findings that derived from the present study could enable the scientists to re-evaluate the potential use of agricultural wastes, especially in the formulation of nutraceuticals

Adenoviral based gene therapy for cancer in human and animals: a review

Adenovirus vector is the most common used vector in clinical gene therapy. The development of adenovirus from the first generation until the helper-dependent adenovirus vector has greatly reduced toxicity and immunogenicity. The helper-dependent adenovirus can also prolong transgene expression. Tissue- or disease-specific approach has been used to improve the specificity of adenoviral vector for cancer gene therapy. This review summarizes some adenoviral gene therapy and targeting approaches available for human cancer as well as animal cancer

Antiviral properties of Berembang Bukit and Kandis Hutan against psedorabies virus in animal cell culture

Tropical rainforest in Malaysia represents and untapped potential source of antiviral compounds. Bioactive compounds in plant species from the same genus as Kandis Hutan such as xanthones, benzophenones, biflavonoids and lupeol had been studied. Eugeniin is an anti-herpesvirus compound which had also been found n Berembang Bukit. This preliminary study was carried out to discover the presence of antiviral properties in Berembang Bukit and Kandis Hutan using different antiviral assays. In this study, MTT cell viability assay was used in addition to microscopic evaluation of pseudorabies virus (PrV)- induced cytopathic effects (CPE) on Vero cells. The cellular toxicity of DMSO was also evaluated. DMSO was less than 10% cytotoxic at concentration of 0.1% to Vero cells and its effect can be negligible. Both plants had demonstrated antiviral properties in thyl acetate and ethanol extracts. From our findings from all three antiviral assays, the ethanol-extracted Kandis Hutan possessed the most promising antiviral properties. Nevertheless, antiviral potential of ethyl acetate and ethanol-extracted Berembang Bukit and ethyl acetate-extracted Kandis Hutan also merit further investigatio

Comparison of clinical signs and mortality rate in murine models infected with herpes virus isolates from captive wildlife

Clinical signs, cellular infectivity and mortality rate of two closely related alpha-herpesvirus from a gaur (UPMV5/05) and a bear(UPMVI9/05) were studied in mice experimentally infected via the nasal route. The grade of viral infection was divided into low (103 and 104 pfu/ml), medium (105 and 106 pfu/ml) and high (107 pfu/ml). Clinically, all animals with the exception of controls, showed respiratory and neurological sign starting from 36 hpi. Although there were a number of differences in the onset time of clinical signs, these were not significant and the types of clinical signs shown were common. Nevertheless, there was a significant difference in the mortality rate, in which isolate UPMVI9/05 from a bear was more pathogenic (83%) than the gaurisolate, UPMV5/05(16.7%). These results strongly suggest that the consequences of host adaptation of herpesviruses may have been derived from interspecies transmission and underscore the need for extreme caution when managing wild or captive animals in close proximity to bovines

Case study of temporal changes in maternal dietary intake and the association with breast milk mineral contents

Minerals play important roles in infant growth and development, even though they only make up to 0.2 g% of the mature breast milk contents. Limited studies examined the association between maternal dietary intake and breast milk mineral contents in a temporal manner. Twenty Malaysian Malay postpartum mothers were recruited by either convenience or snow balling sampling from the urban lower middle income residential areas. Dietary intake of the subjects was obtained by 24-hour recall during each breast milk collection. Adequacy of maternal mineral intake was compared with the latest Recommended Nutrient Intake for Malaysia. Each of the subject provided breast milk samples for three times (T1, T2, and T3) at consecutive 2-week intervals. Breast milk concentrations of selected macro- and micro-minerals, including Ca, P, K, Na, Mg, Fe, Zn, Cu, Mn, Se, I, Cr and Mo were determined by inductively coupled plasma mass spectrometry (ICP-MS). Subjects were aged 31.4 +/- 6.1 years with a majority (60 %) having post-secondary school/college education. While maternal intake of macro-minerals, Ca, P, K and Mg, did not display a significant temporal changes from T1 to T3, the intake of micro-minerals, Cu, Mn and I decreased significantly over time from T1 to T3. Breast milk K, Fe, Zn and Cu concentrations showed a significant decreased with the progression of lactation from transitional (2-3 weeks) to established stage (>8-12 weeks). Significant correlations were established between maternal intake of K, Na, Fe and Se and their respective concentrations in breast milk in the present study. This case study revealed an inadequate maternal intake of several key micro-minerals (Cu, Mn, I) among the postpartum Malay mothers and a decreasing concentrations of certain essential minerals (K, Fe, Zn and Cu) in breast milk with lactation stage

Temporal Changes in Breast Milk Fatty Acids Contents:A Case Study of Malay Breastfeeding Women

The composition of human breast milk changes in the first two months of life, adapting itself to the evolving needs of the growing new-born. Lipids in milk are a source of energy, essential fatty acids (FA), fat-soluble vitamins, and vital bioactive components. Information on breast milk FA of Malaysian lactating women is scarce. Based on convenience sampling, a total of 20 Malay breastfeeding women who fulfilled the inclusion criteria were recruited. Breast milk was collected three times from each subject at consecutive intervals of 2-3 weeks apart. A total of 60 breast milk samples were collected and classified into "transitional milk" (n = 8), "early milk" (n = 26) and "mature milk" (n = 26). All milk samples were air freighted to University of Groningen, Netherlands for analysis. The dominant breast milk FA were oleic acid, constituting 33% of total fatty acids, followed by palmitic acid (26%). Both these FA and the essential FA, linoleic acid (10%) and alpha-linolenic acid (0.4%), showed no significant changes from transitional to mature milk. Breast milk ratio of n-6:n-3 polyunsaturated fatty acids (PUFA) was comparatively high, exceeding 10 throughout the lactation period, suggesting a healthier balance of PUFA intake is needed in pregnancy and at postpartum

Effect of the Presence of Brain-Derived Neurotrophic Factor Val66Met Polymorphism on the Recovery in Patients With Acute Subcortical Stroke

Objective : To investigate the effect of brain-derived neurotrophic factor (BDNF) Valmet polymorphism on the recovery after subcortical stroke, using the modified Rankin Scale (mRS). Methods : Subcortical stroke patients with copies of BDNF Valmet polymorphism (n=7) were compared to their controls (n=7) without a copy of BDNF Valmet polymorphism after matching for initial severity, location and type of stroke. The mRS scores at 1 and 3 months after discharge from the neurorehabilitation unit were compared between the groups. Results : A repeated measures ANOVA for mRS revealed significant interaction between time and group (F(2, 24) =37.2, p<0.001) and a significant effect of time (F(2, 24)=10.8, p<0.001), thereby reflecting significant differences between the Met allele (+) group and the Met allele (-) group. There was a significant difference in mRS scores at 3 months post-discharge between the two groups (p=0.01) although no difference was evident in mRS scores at 1 month post-discharge between the two groups. There were significant improvements between mRS scores on admission and mRS scores at 1 month post-discharge (p=0.02), and between mRS scores at 1 month post-discharge and mRS scores at 3 months post-discharge (p=0.004) in the Met allele (-) group. Conclusion : BDNF Valmet polymorphism may be associated with worse functional outcome in Korean patients with subcortical stroke. Therefore, BDNF Valmet polymorphism should be considered as an important prognostic factor for recovery and responses to rehabilitation therapies after stroke in Korean patients. There is a need for developing different rehabilitation strategies for the population with BDNF Valmet polymorphism. Further studies assessing different outcomes for various functional domains of stroke recovery are needed to clarify the role of BDNF Valmet polymorphism.This research was supported by SK Chemicals Co. through Seoul National University R&DB Foundation (Grant No. 800-2010095).OAIID:oai:osos.snu.ac.kr:snu2013-01/102/0000005165/4SEQ:4PERF_CD:SNU2013-01EVAL_ITEM_CD:102USER_ID:0000005165ADJUST_YN:YEMP_ID:A075663DEPT_CD:801CITE_RATE:0FILENAME:첨부된 내역이 없습니다.DEPT_NM:의학과EMAIL:[email protected]:

Autosomal Dominant Hypocalcemia Caused by an Activating Mutation of the Calcium-Sensing Receptor Gene: The First Case Report in Korea

Hypoparathyroidism is an abnormality of calcium metabolism characterized by low serum levels of parathyroid hormone in spite of hypocalcemia. The causes of hypoparathyroidism are numerous. Activating mutations in the calcium-sensing receptor (CaSR) gene are well-known causes of familial isolated hypoparathyroidism, also known as autosomal dominant hypocalcemia (ADH). Here we describe members of a Korean family with a heterozygous Pro221Leu mutation causing ADH. This case is the first report in Korea