SummaryMutations in RECQL4 helicase are associated with Rothmund-Thomson syndrome (RTS). A subset of RTS patients is predisposed to cancer and is sensitive to DNA damaging agents. The enhanced sensitivity of cells from RTS patients correlates with the accumulation of transcriptionally active nuclear p53. We found that in untreated normal human cells these two nuclear proteins, p53 and RECQL4, instead colocalize in the mitochondrial nucleoids. RECQL4 accumulates in mitochondria in all phases of the cell cycle except S phase and physically interacts with p53 only in the absence of DNA damage. p53-RECQL4 binding leads to the masking of the nuclear localization signal of p53. The N-terminal 84 amino acids of RECQL4 contain a mitochondrial localization signal, which causes the localization of RECQL4-p53 complex to the mitochondria. RECQL4-p53 interaction is disrupted after stress, allowing p53 translocation to the nucleus. In untreated normal cells RECQL4 optimizes de novo replication of mtDNA, which is consequently decreased in fibroblasts from RTS patients. Wild-type RECQL4-complemented RTS cells show relocalization of both RECQL4 and p53 to the mitochondria, loss of p53 activation, restoration of de novo mtDNA replication and resistance to different types of DNA damage. In cells expressing D84 RECQL4, which cannot translocate to mitochondria, all the above functions are compromised. The recruitment of p53 to the sites of de novo mtDNA replication is also regulated by RECQL4. Thus these findings elucidate the mechanism by which p53 is regulated by RECQL4 in unstressed normal cells and also delineates the mitochondrial functions of the helicase.

This study investigated the effects of short and prolonged administration of a yeast beta-glucan on non-specific immune parameters, growth rate and the disease resistance of Asian catfish, Clarias batrachus. Fish fed with a basal diet (control) and test diet (basal diet supplemented with 0.1% glucan) for 1, 2 and 3 weeks were assayed for superoxide production, serum myeloperoxidase (MPO) content, natural haemagglutinin level, complement and lysozyme activities. Fish were weighed at weekly intervals and specific growth rate (SGR, % increase in body weight per day) was determined. After each week, fish were challenged with Aeromonas hydrophila to measure the level of protection. Results showed that glucan administration at 0.1% in feed, significantly (P<0.05) enhanced MPO and lysozyme levels, superoxide production, haemagglutination titre and level of protection against A. hydrophila challenge, irrespective of length of exposure. The alternative complement activity and SGR were not affected by the dietary supplementation of yeast glucan. As glucan feeding at 0.1% for 1 week is able to enhance the non-specific immunity and disease resistance of catfish efficiently, short-term feeding might be used in farmed catfish diets to enhance disease resistance.

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The non-specific immune parameters are useful to determine the health status of fish and to evaluate the immunomodulatory substances for fish farming as markers of pollution and disease resistance. Some of the important parameters, viz. superoxide production by neutrophils through nitroblue tetrazolium (NBT) assay; haemagglutination (HA), haemolysin (HLY) and bacterial agglutination titres; myeloperoxidase (MPO), and lysozyme activities, and alternative complement levels in serum of the juveniles of three Indian major carp species (Cirrhinus mrigala, Catla catla and Labeo rohita) were measured to establish their physiological normal range. A wide variation among the individuals within a species in the ranges of the most of the immune parameters was recorded. Significantly higher levels in the mean values of HA, HLY, bacterial agglutination titres; superoxide production by neutrophils in nitroblue tetrazolium assay; serum MPO and lysozyme activities, i.e. 371.20, 4.60, 18.80, 0.40, 0.62 and 6.55 lg ml )1 , respectively, were obtained in L. rohita except a much lower alternative haemolytic complement activity (29.06 units ml )1 ) compared with the other two species. In most of the parameters, L. rohita showed the highest value, possibly indicating its more natural resistance compared with the other two species.

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The biochemical mechanisms by which RECQL4 and p53 affect mtDNA replication have been elucidated. Resequencing of RTS and LFS patients' mitochondrial genome reveals common mutations indicating similar mechanisms of regulation by RECQL4 and p53.

LBC has been found to be more superior to conventional smears only with respect to lesser number of unsatisfactory smears, but considering the economic implications of LBC, conventional Pap is more feasible in our setting.

Hepatic cell culture on a three-dimensional (3D) matrix or as a hepatosphere appears to be a promising in vitro biomimetic system for liver tissue engineering applications. In this study, we have combined the concept of a 3D scaffold and a spheroid culture to develop an in vitro model to engineer liver tissue for drug screening. We have evaluated the potential of poly(ethylene glycol)-alginate-gelatin (PAG) cryogel matrix for in vitro culture of human liver cell lines. The synthesized cryogel matrix has a flow rate of 7 mL/min and water uptake capacity of 94% that enables easy nutrient transportation in the in vitro cell culture. Young's modulus of 2.4 kPa and viscoelastic property determine the soft and elastic nature of synthesized cryogel. Biocompatibility of PAG cryogel was evaluated through MTT assay of HepG2 and Huh-7 cells on matrices. The proliferation and functionality of the liver cells were enhanced by culturing hepatic cells as spheroids (hepatospheres) on the PAG cryogel using temperature-reversible soluble-insoluble polymer, poly(N-isopropylacrylamide) (PNIPAAm). Pore size of the cryogel above 100 μm modulated spheroid size that can prevent hypoxia condition within the spheroid culture. Both the hepatic cells have shown a significant difference (P < 0.05) in terms of cell number and functionality when cultured with PNIPAAm. After 10 days of culture using 0.05% PNIPAAm, the cell number increased by 11- and 7-fold in case of HepG2 and Huh-7 cells, respectively. Similarly, after 10 days of hepatic spheroids culture on PAG cryogel, the albumin production, urea secretion, and CYP450 activity were significantly higher in case of culture with PNIPAAm. The developed tissue mass on the PAG cryogel in the presence of PNIPAAm possess polarity, which was confirmed using F-actin staining and by presence of intercellular bile canalicular lumen. The developed cryogel matrix supports liver cells proliferation and functionality and therefore can be used for in vitro and in vivo drug testing.