Exposure to Thiodan results in lipofuscin accumulation in hepatocytes of the freshwater catfish Tandanus tandanus

We investigated the effect of time after pulse exposure to 1.0 microg l(-1) endosulfan (applied as Thiodan) on endosulfan residues in the liver and ultrastructural changes in the hepatocytes of the freshwater catfish Tandanus tandanus. Time after exposure did not affect the mean residue level in the liver. After exposure to endosulfan, residues in the liver were 227.47 microg kg(-1) after 1 d and 282.83 microg kg(-1) after 28 d; residues in the bile were 313.97 microg kg(-1) after 1 d and 334.53 microg kg(-1) after 28 d. At the end of 28 d exposure, lipofuscin was present in up to 69% of hepatocytes of fish containing residues of endosulfan, but absent from control fish. There was a statistically significant increase in the percentage of pyknotic nuclei and altered rough endoplasmic reticulum 28 d after exposure. The mean percentage of cells with altered endoplasmic reticulum ranged from 12.93% (Day 1) to 7.50% (Day 28) for control fish, while for exposed fish it increased from 14.30% (Day 1) to 35.00% (Day 28). The mean percentage of cells with pyknotic nuclei increased from 1.1 to 2.1% in control fish and from 3.8 to 9.6% in exposed fish. Other ultrastructural changes included increased ultrastructural heterogeneity, progressive vacuolation and fractionation of rough endoplasmic reticulum, accumulation of lysosomes and residual bodies, intranuclear inclusions and pseudoinclusions, membrane whorls and myelinated bodies. Protracted senescence was one of the main features of endosulfan toxicity to T. tandanus hepatocytes.     

Competitive exclusion and coexistence of universal grammars

Universal grammar (UG) is a list of innate constraints that specify the set of grammars that can be learned by the child during primary language acquisition. UG of the human brain has been shaped by evolution. Evolution requires variation. Hence, we have to postulate and study variation of UG. We investigate evolutionary dynamics and language acquisition in the context of multiple UGs. We provide examples for competitive exclusion and stable coexistence of different UGs. More specific UGs admit fewer candidate grammars, and less specific UGs admit more candidate grammars. We will analyze conditions for more specific UGs to outcompete less specific UGs and vice versa. An interesting finding is that less specific UGs can resist invasion by more specific UGs if learning is more accurate. In other words, accurate learning stabilizes UGs that admit large numbers of candidate grammars.     

Genomic organization and expression profile of the human and mouse <Emphasis Type="Italic">WAVE</Emphasis> gene family

The WAVE gene family, which contains three members, has been shown to play a major role in the actin polymerization and cytoskeleton organization processes. We have identified the WAVE3 gene from Chromosome (Chr) 13q12, as being involved in one of the breakpoints of a t(1:13)(q21:q12) reciprocal translocation, in a patient with ganglioneuroblastoma (Sossey-Alaoui et al. 2002; Oncogene 21: 5967–5974). We have also reported the cloning of the mouse Wave3. During our analysis of the human gene map, we also noted that WAVE2 maps to Chr region lp35-36, which frequently undergoes loss of heterozygosity and deletion in advanced stage neuroblastoma. These data clearly indicate a possible involvement of the WAVE genes in the pathogenesis of neuroblastoma. In this study, we report the complete genomic organization and expression profile of the three human WAVE genes and their mouse orthologs. We show that the WAVE genes have distinctive expression patterns in both adult and fetal human and mouse tissues. We also show a high level of conservation between these genes, in both the nucleotide and protein sequences. We finally show that the genomic structure is highly conserved among these genes and that the mouse Wave genes map to chromosome regions that have synteny in the human genome. The gene content in these syntenic regions is also conserved, suggesting that the WAVE genes are derived from a common ancient ancestor by genome duplication. The genomic characterization and expression analysis of the WAVE genes provide the basis towards understanding the function of these genes. It also provides the first steps towards the development of mouse models for the role of the WAVE genes in actin and cytoskeleton organization in general, and in the development of neuroblastoma in particular.     

The different effects of apoptosis and DNA repair on tumorigenesis

Complex multicellular organisms have evolved mechanisms to ensure that individual cells follow their proper developmental and somatic programs. Tumorigenesis, or uncontrolled cellular proliferation, is caused by somatic mutations to those genetic constraints that normally operate within a tissue. Genes involved in DNA repair and apoptosis are particularly instrumental in safeguarding cells against tumorigenesis. In this paper, we introduce a stochastic framework to analyse the somatic evolution of cancer initiation. Within this model, we study how apoptosis and DNA repair can maintain the transient stability of somatic cells and delay the onset of cancer. Focusing on individual cell lineages, we calculate the waiting time before tumorigenesis in the presence of varying degrees of apoptosis and DNA repair. We find that the loss of DNA repair or the loss of apoptosis both hasten tumorigenesis, but in characteristically different ways.     

Prevalence of TT virus in patients with hepatitis B,C and hepatitis of unknown etiology

Discovery of TT virus in 1997 gave raise to intensive subsequent studies to learn about its structure, features and, what is the most important, about its role in pathogenesis of liver disease. The aim of the work was to analyze prevalence of TTV DNA in patients with diagnosed hepatitis B, C, that of unknown etiology and in healthy blood donors as well. Additionally the divergence of TTV sequence was estimated in selected cases. TTV DNA was detected by PCR technique using specific oligonucleotide primers for coding regions. TT virus has been detected in 25.6% (32/125) HBsAg positive patients and in 23.9% (51/213) HCV infected patients. In healthy blood donors the frequency of TTV was 24.3% (34/140) similarly to that found in HCV and HBV infected patients. The frequency of TTV DNA among patients with hepatitis of unknown etiology was 9.1%. This result was statistically significant lower than in the other groups. When detected sequences have been compared to these from NCBI base the homology result was 71% to 95%, and among different patients and groups of patients identity was 46% to 73%. On the basis of the obtained results it can be concluded that it is very unlikely that TTV coinfection plays any significant role in HCV or HBV infection. The hypothetical role of TTV infection in the etiopathogenesis of cryptogenic chronic hepatitis has not been confirmed. The results obtained in the small group of patients with hepatitis of unknown etiology are not conclusive and should be taken with some precaution. The final conclusion is the TTV coinfection does not contribute to the liver pathology. The divergence of TTV sequences may explain the various frequency of TTV viremia reported by other authors.     

Structural investigation of the binding of nucleotide to phosphoenolpyruvate carboxykinase by NMR

The enzyme phosphoenolpyruvate carboxykinase (PEPCK) catalyzes the reversible conversion of oxalacetate and GTP to phosphoenolpyruvate (PEP), GDP, and CO2. PEPCK from higher organisms is a monomer, specifically requires GTP or ITP, and uses Mn2+ as the activating cation. Currently, there is no crystal structure of GTP-utilizing PEPCKs. The conformation of the bound nucleotide was determined from transferred nuclear Overhauser effects (trnOe) experiments to determine internuclear proton distances. At 600 MHz in the presence of PEPCK, nOe effects were observed between nucleotide protons. Internuclear distances were calculated from the initial rate of the nOe buildup. These distance constraints were used in energy minimization calculations to determine the conformation of PEPCK-bound GTP. The bound nucleotide has the base oriented anti to the C2'-endo(2E) ribose ring conformation. Relaxation rate studies indicate that there is an additional relaxation effect on the C1' proton upon nucleotide binding to PEPCK. Nucleotide binding to PEPCK-Mn2+ was studied by 1H relaxation rate studies, but results were complicated by long dipole-dipole distances and the presence of competing complexes. Modification of PEPCK by iodoacetamido-TEMPO leads to an inactive enzyme that is spin-labeled at cys273. The interaction of TEMPO-PEPCK with GTP allows for the measurement of nuclear distances between GTP and the spin label. The results suggest that cys273 lies near the ribose ring of the bound nucleotide, but it is too far to be implicated in direct hydrogen bonding interactions consistent with previous results [Makinen, A. L., and Nowak, T. J. Biol. Chem. (1989) 264, 12148], suggesting that cys273 does not actively participate in catalysis. Modification of PEPCK with several cysteine specific modifying agents causes no change in the ability of the enzyme to bind nucleotide as monitored by fluorescence quenching. A correlation between the size of the modifying agent and the maximal observed quenching upon saturation of the enzyme with nucleotide is observed. This suggests a mechanism for inactivation of PEPCK by cysteine modification due to inhibition of a dynamic motion that may occur upon nucleotide binding.