Effects of H pylori infection on gap-junctional intercellular communication and proliferation of gastric epithelial cells in vitro

AIM： To explore the effects of H pylori infection on gap-junctional intercellular communication （GJIC） and proliferation of gastric epithelial cells in vitro. METHODS： A human gastric epithelial cell line （SGC- 7901） cultured on coverslips was exposed overnight to intact H pylori （CagA^＋ or CagA^- strains） and sonicated extracts, respectively. GJIC between the cells was detected by fluorescence redistribution after photobleaching （FRAP） technique. Proliferation of SGC cells was determined by methylthiazolyl tetrazolium （MTT） assay. RESULTS： When compared with control in which cells were cultured with simple medium alone, both CagA^＋ and CagA^- H pylori isolates could inhibit GJIC （CagA^＋： F = 57.98, P 〈 0.01; CagA^-： F = 29.59, P 〈 0.01） and proliferation （CagA^＋： F = 42.65, P 〈 0.01; CagA^-： F = 58.14, P 〈 0.01） of SGC-7901 cells. Compared with CagA^- strains, CagA^＋ H pylori more significantly downregulated GJIC of gastric cells （intact Hpylori： t = 13.86, P 〈 0.01; sonicated extracts： t = 11.87, P 〈 0.01） and inhibited proliferation gastric cells to a lesser extent in vitro （intact H pylori： t = 3.06, P 〈 0.05; sonicated extracts： t = 3.94, P 〈 0.01）. CONCLUSION： Compared with CagA^- H pylori strains, CagA^＋ strains down-regulate GJIC of gastric epithelial cells more significantly and inhibit proliferation of gastric cells to a lesser extent in vitro. H pylori, especially CagA^＋ strains, may play an important role in gastric carcinogenesis.

Evaluation of <i>In-Vitro</i>Anti-Tuberculosis Activity of <i>Tetrapleura tetraptera</i>Crude and Fractions on Multidrug Resistant <i>Mycobacterium tuberculosis</i>

The management, control and elimination of tuberculosis (TB) have been difficult with the advent of HIV and cases of multidrug resistant (MDR-TB) tuberculosis. The cases of multidrug resistance to rifampicin and isoniazid pose greater challenges on first line and second line drugs to eliminate TB. The study is aimed at establishing anti-tuberculosis activity of Tetrapleura tetraptera against Mycobacterium tuberculosis and MDR-TB and the phytochemical present. The leaves of Tetrapleura tetraptera were collected, weighed, dried and pulverized to powder. The pulverized leaves of Tetrapleura tetraptera were subjected to 70% methanol extraction and screened for phytochemical. The crude extract was further purified into fractions using silica gel and thin layer chromatography techniques. M. tuberculosis and MDR-TB were obtained from positive acid fast bacilli sputa of TB patients and confirmed using GeneXpert to differentiate genotypic drug susceptible M. tuberculosis and MDR-TB. The sputa were digested using sodium hydroxide-cysteine technique and cultured in Middlebrook 7H9. The crude extract and fractions were screened for anti-tuberculosis activity using tetrazolium microtitre plate assay. The results showed that Tetrapleura tetraptera crude had activities against M. tuberculosis at 7.4 ± 0 mg/ml and 27.5 ± 0 mg/ml for MDR-TB. One of the fractions inhibited the growth of M. tuberculosis at 0.24 ± 0 mg/ml and MDR-TB at 0.89 ± 0 mg/ml. The phytochemical screened includes tannins, alkaloids, saponins, flavonoids, phenols and resins. T. tetraptra possesses anti-tuberculosis potential at low concentration on MDR-TB and can be a lead compound in drug development for the treatment of tuberculosis and multidrug resistant tuberculosis.

Morphological and behavioural variation in CNS innate defence cell microglia is development and age sensitive

Aim:Microglia,the innate defence cells in central nervous system(CNS),alters their shapes and function with age.We observed and identified these morphological changes and functional association throughout the developmental gradient until adulthood in rat brain.Methods:Early and late embryonic stages,neonates and adult brains of albino rats were sectioned for routine Haematoxylin Eosin(HE)staining and specialized silver-gold staining to show distribution and morphological variation in situ.Isolated microglia from different age groups was subjected to scanning electron microscopy(SEM)for observing ultrastructural shapes of microglial cells.The Viability of isolated cells was measured by trypan blue staining and their cellular identity by immuno-staining for CD11b.Finally,phagocytic limitations of the cells in normally developing brain were assessed by carbon particle ingestion and oxidative burst through nitroblue tetrazolium assay to investigate microglial age-sensitivity behavioural response.Results:HE staining spotted overall cellular distribution in the brain and cells with monocytic appearance among the other CNS cells.On contrary,silver-gold staining showed variable morphologies of microglia in various age groups and also showed the appearance of ramified microglia in adult.Nearly 90%of isolated cells were viable and positive for CD11b.SEM showed variable shapes of amoeboid and ramified forms.Immunofluoresence confirms microglial identity.Functionally,microglia showed an age dependent baseline phagocytic capacity in normal condition which changes with developmental phase and age with most active phagocytic behaviour around perinatal phase.Conclusion:In normally developing brains,microglia shows variability in morphology and baseline phagocytic activity that changes with age.These results may represent the normal physiology of CNS development and function.

Protective effects of Amaranthus hybridus against aflatoxin B1 and fumonisin B1-induced genotoxicity in H4IIE-luc cells

Aim:Protective effects of aqueous extract of Amaranthus hybridus against afl atoxin B1(AFB_(1))and/or fumonisin B1(FB_(1))on the H4IIE-luc cell line were determined by use of the methyl thiazol tetrazolium viability assay and disruption of DNA integrity.Methods:H4IIE-luc cells were incubated with different concentrations of AFB_(1) and/or FB_(1) for 24 and 48 h with or without aqueous extract of A.hybridus.Results:AFB_(1) decreased the viability of cells after 24 and 48 h of exposure.EC_(50)values for AFB_(1) were 10.5 and 1.8μmol/L for the two periods,respectively.When the 48 h exposure to mycotoxin repeated with a pre-treatment of 20 and 40μg/mL extract of A.hybridus,the EC_(50)changed to 3.88 and 7.67μmol/L,respectively.H4IIE-luc cells exposed to FB_(1) for 24 h responded more than those incubated for 48 h.Cells treated with a combination of AFB_(1) and FB_(1) were less viable with a signifi cant decrease in the greater concentration.The mixture of AFB_(1) and FB_(1) resulted in a signifi cant threat to H4IIE-luc as indicated by the absence or appearance of new bands in random amplifi ed polymorphic DNA analysis,which demonstrated damage to DNA.The protective effects were probably due to greater content of total phenolics,carotenoids,β-carotene,folic-,linolenic-,linoleic and palmitic acids,as well as calcium,magnesium,iron,zinc,and selenium observed in the extract.Conclusion:Exposure to 40μg/mL of extract of A.hybridus protected cells from damage to DNA by stabilizing DNA.