Rotavirus induces apoptosis in fully differentiated human intestinal Caco-2 cells.

Rotaviruses, which are the main cause of viral gastroenteritis in young children, induce structural and functional damages in infected mature enterocytes of the small intestine. To investigate a relationship between rotavirus infection and cell death by apoptosis, we used the human intestinal Caco-2 cell line. We demonstrated by several methods including TUNEL and ELISA detection of cytoplasmic histone-associated DNA fragments that the infection of fully differentiated Caco-2 cells by the RRV rotavirus strain induces apoptosis. Rotavirus infection leads to the loss of mitochondrial membrane potential and the release of cytochrome C from mitochondria. We showed that rotavirus-induced apoptosis was dependent of the multiplicity of infection and increased with time from 4 h to 24 h of infection. Flow cytometric analysis showed that DNA fragmentation occurs in productively infected cells, suggesting that rotavirus induces apoptosis by a direct mechanism. We also demonstrated that non-replicative RRV particles are not sufficient to induce apoptosis and viral gene expression seems required. Intracellular calcium plays a role in RRV-induced apoptosis because treatment with an intracellular calcium ion chelator (BAPTA-AM) partially inhibited apoptosis.

A cyclic AMP protein kinase A-dependent mechanism by which rotavirus impairs the expression and enzyme activity of brush border-associated sucrase-isomaltase in differentiated intestinal Caco-2 cells.

We undertook a study of the mechanism by which rhesus monkey rotavirus (RRV) impairs the expression and enzyme activity of brush border-associated sucrase isomaltase (SI) in cultured, human, fully differentiated, intestinal Caco-2 cells. We provide evidence that the RRV-induced defects in the expression and enzyme activity of SI are not related to the previously observed, RRV-induced, Ca2+ -dependent, disassembly of the F-actin cytoskeleton. This conclusion is based on the facts that: (i) the intracellular Ca2+ blocker, BAPTA/AM, which antagonizes the RRV-induced increase in [Ca2+](i), fails to inhibit the RRV-induced decrease in SI expression and enzyme activity; and (ii) Jasplakinolide (JAS) treatment, known to stabilize actin filaments, had no effect on the RRV-induced decrease in SI expression. Results reported here demonstrate that the RRV-induced impairment in the expression and enzyme activity of brush border-associated SI results from a hitherto unknown mechanism involving PKA signalling. This conclusion is based on the observations that (i) intracellular cAMP was increased in RRV-infected cells and (ii) treatment of RRV-infected cells with PKA blockers resulted in the reappearance of apical SI expression, accompanied by the restoration of the enzyme activity at the brush border. In addition, in RRV-infected cells a twofold increase of phosphorylated form of cytokeratin 18 was observed after immunopurification and Western Blot analysis, which was antagonized by exposing the RRV-infected cells to the PKA blockers.

Síntese de tiossemicarbazonas derivadas de 2-(fenoxi)acetaldeídos com potencial atividade antiviral / Synthesis of some thiosemicarbazones of 2-(phenoxy)acetaldehydes as potential antiviral agents

As tiossemicarbazonas constituem classe de compostos que têm apresentado amplo espectro de ação, englobando atividades antibeoplásica, antiinflamatória, tuberculostática e antiviral, inclusive anti-HIV. Diante da potencial atividade antiviral das tiossemicarbazonas, planejamos sintetizar nova série de compostos a partir da tiossemicarbazida e fenoxiacetaldeídos diversamente substituídos. Os produtos obtidos tiveram sua estrutura química comprovada por meio de métodos espectroscópicos no infravermelho e ressonância magnética nuclear de hidrogênio e microanálise. Cinco compostos foram testados visando encontrar possível atividade antiviral. Nos ensaios utilizaram-se culturas de células contínuas VERO e L929 infectadas pelo vírus herpes humano tipo I, da vaccínia, poliomielítico tipo I e da estomatite vesiculosa...