Involvement of pro-apoptotic and anti-apoptotic factors in the early development of the human pituitary gland

The spatial and temporal pattern of appearance of pro-apoptotic caspase-3 and p53 proteins, and anti-apoptotic bcl-2 protein was investigated in the developing pituitary gland of 6 human embryos 5-8-weeks old, using morphological and immunohistochemical techniques. Their dynamic appearance was analyzed in the Rathke's pouch (future adenohypophysis), mesenchyme, and in the developing neurohypophysis. In the 5th and 6th week, caspase-3 positive cells appeared in the Rathke's pouch (5%) and stalk (11%), in the mesenchyme, but not in the neurohypophysis. In the 6th and 7th week, apoptotic cells were more numerous in the caudal part of the Rathke's pouch due to its separation from the oral epithelium. Pro-apoptotic p53 protein was detected in all parts of the pituitary gland throughout the investigated period. Nuclear condensations characterized cells positive to caspase-3 and p53 proteins. Apoptotic cells displayed condensations of nuclear chromatin on an ultrastructural level as well. While caspase-3 dependent pathway of cell death participated in morphogenesis of the adenohypophysis and associated connective tissue, p53-mediated apoptosis most likely participates in morphogenesis of all parts of the gland, including neurohypophysis. The anti-apoptotic bcl-2 protein was also detected in all parts of the developing gland. With advancing development, the positivity to bcl-2 protein increased in the cells of the adenohypophysis, while it decreased in the neurohypophysis. Bcl-2 protein probably prevented cell death in all parts of the gland and enhanced cell differentiation. The described pattern of appearance of the investigated pro-apoptotic and anti-apoptotic factors might be important for normal morphogenesis and function of the pituitary gland.     

The occurrence of c-myc, p53 and Bcl-2 family proteins in the early phase of development of duodenal epithelium

In last few years, numerous groups of proteins participating in the regulation of cell proliferation, differentiation and death during ontogenesis have been described. In this study we compared the occurrence of Bcl-2, p53 and myc protein families with the level of proliferative activity and apoptosis during development of duodenal epithelium. Paraffin embedded tissues of eight human embryos and foetuses aged from the 6th-18th week of IUD were used. For the detection of apoptotic cells the TUNEL method was performed, the proliferative marker PCNA and all the proteins studied were detected by means of indirect three-step immunohistochemical method. In the 6th and 8th week of intrauterine development we observed isolated TUNEL positive epithelial cells only and this was accompanied by the disperse presence of PCNA as well as by all the studied proteins: Bcl-2, Bax, Bcl-XL, c-myc, N-myc, p53, p63 and p73. In the early foetal period of duodenal development we registered changes in PCNA and TUNEL positivity in accordance with the constitution of the stem cell pool on base of villi, where more numerous Bcl-2 positive cells were also found. The separation of primitive crypts and villi was not accompanied by any differences in distribution of Bax, Bcl-XL, c-myc, N-myc, p63 and p73 proteins between those compartments: all the studied proteins showed dispersed character. P53 rapidly decreased in this period. In the 18th week of intrauterine development the balance between proliferation in crypts and apoptosis of villi epithelium was well established and no p53 positive cells were found. In the presence of Bcl-2, Bax, Bcl-XL, p63 and p73 we did not find any dramatic changes. The myc proteins were restricted within the epithelium of the Lieberkuhn crypts only.     

Effects of specific and prolonged expression of zebrafish growth factors,Fgf2 and Lif in primordial germ cells in vivo

Primordial germ cells (PGCs), specified early in development, proliferate and migrate to the developing gonad before sexual differentiation occurs in the embryo and eventually give rise to spermatogonia or oogonia. In this study, we discovered that nanos3 3′UTR, a common method used to label PGCs, not only directed PGC-specific expression of DsRed but also prolonged this expression up to 26 days post fertilization (dpf) when DsRed-nanos3 3′UTR hybrid mRNAs were introduced into 1- to 2-cell-stage embryos. As such, we employed this knowledge to express zebrafish leukemia inhibitory factor (Lif), basic fibroblast growth factor (Fgf2) and bone morphogenetic protein 4 (Bmp4) in the PGCs and evaluate their effects on PGC development in vivo for over a period of 3 weeks. The results show that expression of Fgf2 significantly increased PGC number at 14- and 21-dpf while Bmp4 resulted in severe ventralization and death of the embryos by 3 days. Expression of Lif resulted in a significant disruption of PGC migration. Mopholino knockdown experiments indicated that Lif illicited its effect on PGC migration through Lif receptor a (Lifra) but not Lifrb. The general approach described in this study could be used to achieve prolonged PGC-specific expression of other proteins to investigate their roles in germ cell and gonad development. The results also indicate that zebrafish PGCs have a mechanism to stabilize and prolong the expression of mRNA that carries nanos3 3′UTR. Understanding this mechanism may make it possible to achieve prolonged RNA expression in other cell types.     

An electron microscopic study of early gonadogenesis in the hermaphroditic appendicularian Oikopleura gracilis (Tunicata,Oikopleuridae)

The postembryonic development of the gonad in the hermaphroditic appendicularian O. gracilis was studied using transmission electron microscopy. The primordial germ cells were detected first in 10-h-old larvae and represent migrating primordial germ syncytium (mPGS) localized in the hemocoel of the tail/trunk junction and several haemocoel areas of the digestive compartment. The mPGS consisted of primordial germ nuclei (PGN) 2 μm in diameter, and elongate somatic-line nuclei 1.8 μm in diameter. In 12.5-h-old juveniles the gonad primordium 40 × 90 μm in size, was separated by a narrow space of haemocoel between the gut and the epidermis of the reproductive compartment. The gonad primordium consisted of the central syncytial part of primordial germ nuclei (PGN), enclosing a single layer of somatic epithelium. In 3-day-old juveniles, the gonad was differentiated into testis and ovary. The testis, 400 × 550 μm in size, is a syncytium of spermatogonial nuclei, covered by a single layer of somatic epithelium. The ovaries, 350 × 850 μm in size, consist of a syncytium with nurse nuclei and meiotic nuclei. The hermaphroditic gonad originates from extragonadal mPGS. Early gonadogenesis in appendicularians has ultrastructural features in common with early gonadogenesis in ascidians.     

A Study on the Anticarcinogenic Effects of Calcium Fructoborate

Evidences about the preventive and therapeutic effects of boron compounds on cancer have been increasing in the last years. Although calcium fructoborate (CaFB) is used as a nutritional supplement, data about its preventive and therapeutic effects on neoplastic transformations are limited. In the present study, the various concentrations of CaFB were applied to the MDA-MB-231 metastatic breast cancer cell line. First, we examined the cytotoxic effect and IC₅₀ value of CaFB by MTT assay. For the evaluation of the DNA damage, apoptosis and metastatic potential, expression levels of ATM, pATM, PARP, p53, p-p53, caspase-3, caspase-9, and VEGF were investigated by using immunoblotting and immunohistochemical methods. Cell viability was significantly reduced at 50 μM CaFB treatment. pATM, p-p53, and caspase-9 levels increased significantly in all groups; furthermore, there was approximately 12.5-, 2.4-, and 10.7-fold increase, respectively, for 100 μM CaFB treatment. ATM and p53 levels did not change with CaFB treatment, but PARP levels significantly 2.5-fold decreased. While VEGF immunoreactivity decreased in all groups, significant increase in caspase-3 immunoreactivity was observed only in the group treated with 50 μM CaFB (p < 0,001). Our results imply that CaFB may have therapeutic potential as well as preventive benefits in cancer.     

Characterization of the cell death induced by cadmium in HaCaT and C6 cell lines

Cell death resulting from cadmium (Cd) intoxication has been confirmed to induce both necrosis and apoptosis. The ratio between both types of cell death is dose- and cell-type-dependent. This study used the human keratinocytes HaCaT expressing a mutated p53 and the rat glial cells C6 expressing a wild p53 as models to characterize Cd-induced apoptosis, using sub-lethal and lethal doses. At these concentrations, features of apoptosis were observed 24 h after C6 cell treatment: apoptotic DNA fragmentation and caspase-9 activation, whereas Cd did not induce caspase-3. In HaCaT, Cd did not induce apoptotic DNA fragmentation or caspase-9 and -3 activation. The results also showed that the inhibition of p53 led to a resistance of the C6 cells to 20 µm Cd, decreased the apoptosis and increased the metallothioneins in these cells. p53 restoration increased the sensitivity of HaCaT cells to Cd but did not affect the MT expression. The results suggest that Cd induced apoptosis in C6 cells but a non-apoptotic cellular death in HaCaT cells.     

Role of thymidine phosphorylase in Fas-induced apoptosis

Thymidine phosphorylase (TP) has chemotactic and angiogenic activity in vitro, and it promotes tumor growth and inhibits apoptosis in vivo. It plays a key role in the invasiveness and metastasis of TP-expressing solid tumors. KB/TP cells transfected with a TP cDNA have been shown to be resistant to hypoxia-induced apoptosis, suggesting that TP has effects on tumor growth and cell death independent of its effects on angiogenesis. However, the mechanisms of cell death inhibition by TP are unknown. In the present study, we demonstrate that caspase-8 is cleaved in control transfectant KB cells early on during Fas-induced apoptosis. Caspase-8 activation leads to the loss of mitochondrial membrane potential, followed by the release of cytochrome c, the activation of caspase-3, and apoptosis. In contrast, Fas-induced caspase-8 cleavage is inhibited in KB/TP cells, which lead to inhibition of the downstream apoptotic cascade and inhibition of apoptosis. These findings indicate that TP plays an important role in intracellular apoptotic signal transduction in the Fas-induced apoptotic pathway. Therefore, inhibition of TP may suppress the progression of TP-overexpressing solid tumors by inducing apoptosis.     

The role of bronchial epithelial cell apoptosis in the pathogenesis of COPD

There is an increased airway inflammation in the pathogenesis of chronic obstructive pulmonary disease (COPD), and it has been suggested that there may also be problem in the apoptosis and renewal of cells. However, there are limited human airway cell studies, in particular those from larger airways such as bronchi. We cultured primary human bronchial epithelial cells (HBECs) from bronchial explants of smokers (n = 6) without COPD and smokers with COPD (n = 8). Apoptosis was studied by fluorescence activated cell sorting. qRT-PCR was used to assess mRNA expression for proteins involving apoptosis including p21^CIP1/WAF1, p53, caspase-8 and caspase-9. Although there was no difference in the rate of viable cells between cells from smokers and COPDs, the level of early apoptotic cells was significantly increased in COPD cells [mean ± standard error of mean (SEM) = 4.86 ± 3.2 %, p = 0.015] as compared to smokers (mean ± SEM = 2.71 ± 1.62 %). In contrast, the rate of late apoptotic cells was significantly decreased in COPD cells (mean ± SEM = 9.82 ± 5.71 %) comparing to smokers (mean ± SEM = 15.21 ± 5.08 %, p = 0.003). Although expression of mRNA for p21^CIP1/WAF1 and caspase-9 was similar in both groups, p53 and caspase-8 mRNA expression was significantly greater in COPD cells. These findings suggest that HBEC apoptosis is increased in COPD, and that this involves p53 and caspase-8 pathways.     

Betaine Protects Against Rotenone-Induced Neurotoxicity in PC12 Cells

Rotenone is an inhibitor of mitochondrial complex I-induced neurotoxicity in PC12 cells and has been widely studied to elucidate the pathogenesis of Parkinson’s disease. We investigated the neuroprotective effects of betaine on rotenone-induced neurotoxicity in PC12 cells. Betaine inhibited rotenone-induced apoptosis in a dose-dependent manner, with cell viability increasing from 50 % with rotenone treatment alone to 71 % with rotenone plus 100-μM betaine treatment. Flow cytometric analysis demonstrated cell death in the rotenone-treated cells to be over 50 %; the number of live cells increased with betaine pretreatment. Betaine pretreatment of PC12 cells attenuated rotenone-mediated mitochondrial dysfunction, including nuclear fragmentation, ATP depletion, mitochondrial membrane depolarization, caspase-3/7 activation, and reactive oxygen species production. Western blots demonstrated activation of caspase-3 and caspase-9, and their increased expression levels in rotenone-treated cells; betaine decreased caspase-3 and caspase-9 expression levels and suppressed their activation. Together, these results suggest that betaine may serve as a neuroprotective agent in the treatment of neurodegenerative diseases.     

microRNA-210 is upregulated in hypoxic cardiomyocytes through Akt- and p53-dependent pathways and exerts cytoprotective effects

microRNA-210 (miR-210) is upregulated in hypoxia, but its function in cardiomyocytes and its regulation in response to hypoxia are not well characterized. The purpose of this study was to identify upstream regulators of miR-210, as well as to characterize miR-210's function in cardiomyocytes. We first showed miR-210 is upregulated through both hypoxia-inducible factor (HIF)-dependent and -independent pathways, since aryl hydrocarbon nuclear translocator (ARNT) knockout mouse embryonic fibroblasts (MEF), lacking intact HIF signaling, still displayed increased miR-210 levels in hypoxia. To determine the mechanism for HIF-independent regulation of miR-210, we focused on p53 and protein kinase B (Akt). Overexpression of p53 in wild-type MEFs induced miR-210, whereas p53 overexpression in ARNT knockout MEFs did not, suggesting p53 regulates miR-210 in a HIF-dependent mechanism. Akt inhibition reduced miR-210 induction by hypoxia, whereas Akt overexpression increased miR-210 levels in both wild-type and ARNT knockout MEFs, indicating Akt regulation of miR-210 is HIF-independent. We then studied the effects of miR-210 in cardiomyocytes. Overexpression of miR-210 reduced cell death in response to oxidative stress and reduced reactive oxygen species (ROS) production both at baseline and after treatment with antimycin A. Furthermore, downregulation of miR-210 increased ROS after hypoxia-reoxygenation. To determine a mechanism for the cytoprotective effects of miR-210, we focused on the predicted target, apoptosis-inducing factor, mitochondrion-associated 3 (AIFM3), known to induce cell death. Although miR-210 reduced AIFM3 levels, overexpression of AIFM3 in the presence of miR-210 overexpression did not reduce cellular viability either at baseline or after hydrogen peroxide treatment, suggesting AIFM3 does not mediate miR-210's cytoprotective effects. Furthermore, HIF-3α, a negative regulator of HIF signaling, is targeted by miR-210, but miR-210 does not modulate HIF activity. In conclusion, we demonstrate a novel role for p53 and Akt in regulating miR-210 and demonstrate that, in cardiomyocytes, miR-210 exerts cytoprotective effects, potentially by reducing mitochondrial ROS production.     

TP53INP1, a tumor suppressor, interacts with LC3 and ATG8-family proteins through the LC3-interacting region (LIR) and promotes autophagy-dependent cell death

TP53INP1 (tumor protein 53-induced nuclear protein 1) is a tumor suppressor, whose expression is downregulated in cancers from different organs. It was described as a p53 target gene involved in cell death, cell-cycle arrest and cellular migration. In this work, we show that TP53INP1 is also able to interact with ATG8-family proteins and to induce autophagy-dependent cell death. In agreement with this finding, we observe that TP53INP1, which is mainly nuclear, relocalizes in autophagosomes during autophagy where it is eventually degraded. TP53INP1-LC3 interaction occurs via a functional LC3-interacting region (LIR). Inactivating mutations of this sequence abolish TP53INP1-LC3 interaction, relocalize TP53INP1 in autophagosomes and decrease TP53INP1 ability to trigger cell death. Interestingly, TP53INP1 binds to ATG8-family proteins with higher affinity than p62, suggesting that it could partially displace p62 from autophagosomes, modifying thereby their composition. Moreover, silencing the expression of autophagy related genes (ATG5 or Beclin-1) or inhibiting caspase activity significantly decreases cell death induced by TP53INP1. These data indicate that cell death observed after TP53INP1-LC3 interaction depends on both autophagy and caspase activity. We conclude that TP53INP1 could act as a tumor suppressor by inducing cell death by caspase-dependent autophagy.     

Paeonol exhibits anti-tumor effects by apoptotic and anti-inflammatory activities in 7,12-dimethylbenz(a)anthracene induced oral carcinogenesis

We investigated the preventive potential of paeonol on 7,12-dimethylbenz(a)anthracene (DMBA) induced oral carcinogenesis. Oral tumors were developed in the buccal pouches of Syrian golden hamsters using topical application of 0.5% DMBA three times/week for 10 weeks. DMBA treated hamsters developed hyperplasia, dysplasia and well-differentiated squamous cell carcinoma. The animals also exhibited increased lipid oxidation, decreased antioxidant status and altered levels of detoxification agents. Paeonol treatment of DMBA treated hamsters for 14 weeks decreased tumor incidence, volume and burden Paeonol treatment also increased antioxidant activity and decreased lipid oxidation to near normal levels. Histomorphology and the expression patterns of mutant p53, cyclo-oxygenase (COX-2) and caspase-9 were investigated in the oral buccal mucosa. Paeonol exhibited protective effects against DMBA induced oral carcinogenesis owing to its antitumor, antioxidant, anti-inflammatory and apoptosis inducing properties.     

Coactivation of STAT and Ras is required for germ cell proliferation and invasive migration in Drosophila

Primordial germ cells (PGCs) undergo proliferation, invasion, guided migration, and aggregation to form the gonad. Here we show that in Drosophila, the receptor tyrosine kinase Torso activates both STAT and Ras during the early phase of PGC development, and coactivation of STAT and Ras is required for PGC proliferation and invasive migration. Embryos mutant for stat92E or Ras1 have fewer PGCs, and these cells migrate slowly, errantly, and fail to coalesce. Conversely, overactivation of these molecules causes supernumerary PGCs, their premature transit through the gut epithelium, and ectopic colonization. A requirement for RTK in Drosophila PGC development is analogous to the mouse, in which the RTK c-kit is required, suggesting a conserved molecular mechanism governing PGC behavior in flies and mammals.     

Association between the TP53 polymorphisms and lung cancer risk: a meta-analysis

The previous published data on the association between TP53 codon 72, intron 6, and intron 3 16 bp polymorphisms and lung cancer risk remained controversial. This meta-analysis of literatures was performed to derive a more precise estimation of the relationship. 38 publications with 51 studies were selected for this meta-analysis, including 17,337 cases and 16,127 controls for TP53 codon 72 (from 43 studies), 2,201 cases and 2,399 controls for TP53 intron 6 (from four studies), and 4,322 cases and 4,558 controls for TP53 intron 3 16 bp (from four studies). When all the eligible studies were pooled into the meta-analysis of codon 72 polymorphism, there was significant association between lung cancer risk and codon 72 polymorphism in any genetic model (dominant model: OR = 1.13, 95 % CI 1.05–1.21; recessive model: OR = 1.14, 95 % CI 1.02–1.27; additive model: OR = 1.19, 95 % CI 1.05–1.33). In the subgroup analysis by ethnicity, histological type, source of control, and smoking status, significantly increased risks were observed in subgroups such as Asians, Caucasians, lung squamous cell carcinoma patients for Asians, population-based study, hospital-based study, non-smokers, and smokers. When all the eligible studies were pooled into the meta-analysis of intron 6 polymorphism, there was significant association between lung cancer risk and intron 6 polymorphism in dominant model (OR = 1.27, 95 % CI 1.11–1.44). When all the eligible studies were pooled into the meta-analysis of intron 3 16 bp polymorphism, there was significant association between lung cancer risk and intron 3 16 bp polymorphism in dominant model (OR = 1.12, 95 % CI 1.02–1.23) and additive model (OR = 1.41, 95 % CI 1.04–1.90). Additionally, when one study was deleted in the sensitive analysis, the results of TP53 intron 3 16 bp duplication polymorphism were changed in the dominant model (OR = 1.11, 95 % CI 0.87–1.42) and additive model (OR = 1.01, 95 % CI 0.65–1.56). In summary, this meta-analysis indicates that codon 72 and intron 6 polymorphisms show an increased lung cancer risk. A study with the larger sample size is needed to further evaluated gene-environment interaction on TP53 codon 72, intron 6, and intron 3 16 bp polymorphisms and lung cancer risk.     

A Novel Piggybac Transposon Inducible Expression System Identifies a Role for Akt Signalling in Primordial Germ Cell Migration

In this work, we describe a single piggyBac transposon system containing both a tet-activator and a doxycycline-inducible expression cassette. We demonstrate that a gene product can be conditionally expressed from the integrated transposon and a second gene can be simultaneously targeted by a short hairpin RNA contained within the transposon, both in vivo and in mammalian and avian cell lines. We applied this system to stably modify chicken primordial germ cell (PGC) lines in vitro and induce a reporter gene at specific developmental stages after injection of the transposon-modified germ cells into chicken embryos. We used this vector to express a constitutively-active AKT molecule during PGC migration to the forming gonad. We found that PGC migration was retarded and cells could not colonise the forming gonad. Correct levels of AKT activation are thus essential for germ cell migration during early embryonic development.     

Endoplasmic reticulum stress,autophagic and apoptotic cell death,and immune activation by a natural triterpenoid in human prostate cancer cells

Though the current therapies are effective at clearing an early stage prostate cancer, they often fail to treat late-stage metastatic disease. We aimed to investigate the molecular mechanisms underlying the anticancer effects of a natural triterpenoid, ganoderic acid DM (GA-DM), on two human prostate cancer cell lines: the androgen-independent prostate carcinoma (PC-3), and androgen-sensitive prostate adenocarcinoma (LNCaP). Cell viability assay showed that GA-DM was relatively more toxic to LNCaP cells than to PC-3 cells (IC₅₀s ranged 45-55 µM for PC-3, and 20-25 µM for LNCaP), which may have occurred due to differential expression of p53. Hoechst DNA staining confirmed detectable nuclear fragmentation in both cell lines irrespective of the p53 status. GA-DM treatment decreased Bcl-2 proteins while it upregulated apoptotic Bax and autophagic Beclin-1, Atg5, and LC-3 molecules, and caused an induction of both early and late events of apoptotic cell death. Biochemical analyses of GA-DM-treated prostate cancer cells demonstrated that caspase-3 cleavage was notable in GA-DM-treated PC-3 cells. Interestingly, GA-DM treatment altered cell cycle progression in the S phase with a significant growth arrest in the G2 checkpoint and enhanced CD4 ⁺ T cell recognition of prostate tumor cells. Mechanistic study of GA-DM-treated prostate cancer cells further demonstrated that calpain activation and endoplasmic reticulum stress contributed to cell death. These findings suggest that GA-DM is a candidate for future drug design for prostate cancer as it activates multiple pathways of cell death and immune recognition.     

Kinetics of Apoptosis and Expression of Apoptosis-Related Proteins in Rat CA3 Hippocampus Cells After Experimental Diffuse Brain Injury

The present study examined kinetics of apoptosis and expression of apoptosis-related proteins Bcl-2, Bax, and caspase-3 in the CA3 hippocampus cells after diffuse brain injury (DBI) induced experimentally in rats. Percentage of apoptotic cells and expressions of above proteins were examined by flow cytometry and immunohistochemistry. Substantial neuronal apoptosis was documented in the CA3 hippocampus cells after DBI (22.26 ± 2.97 % at 72 h after DBI vs. 2.92 ± 0.88 % in sham-operated animals). Expression of Bc1-2 decreased, while expression of Bax and caspase-3 increased after DBI, with caspase-3 expression peaking after that of Bax (72 vs. 48 h, respectively). Further, the Bc1-2/Bax expression ratio decreased prior to increase of caspase-3 expression. In conclusion, cell apoptosis and altered expressions of Bcl-2, Bax, and caspase-3 are present in the CA3 region of hippocampus after experimental DBI. Changes in the Bc1-2/Bax expression ratio may facilitate activation of caspase-3 and aggravate neuronal apoptosis after brain injury.     

Decreased PGC1-α levels and increased apoptotic protein signaling are associated with the maladaptive cardiac hypertrophy in hyperthyroidism

Hyperthyroidism can lead to the activation of proteins which are associated with inflammation, apoptosis, hypertrophy, and heart failure. This study aimed to explore the inflammatory and apoptotic proteins involved in the hyperthyroidism-induced cardiac hypertrophy establishment. Male Wistar rats were divided into control and hyperthyroid (12 mg/L L-thyroxine, in drinking water for 28 days) groups. The expression of inflammatory and apoptotic signaling proteins was quantified in the left ventricle by Western blot. Hyperthyroidism was confirmed by evaluation of T3 and T4 levels, as well as cardiac hypertrophy development. There was no change in the expression of HSP70, HIF1-α, TNF-α, MyD88, p-NFκB, NFκB, p-p38, and p38. Reduced expression of p53 and PGC1-α was associated with increased TLR4 and decreased IL-10 expression. Decreased Bcl-2 expression and increased Bax/Bcl-2 ratio were also observed. The results suggest that reduced PGC1-α and IL-10, and elevated TLR4 proteins expression could be involved with the diminished mitochondrial biogenesis and anti-inflammatory response, as well as cell death signaling, in the establishment of hyperthyroidism-induced maladaptive cardiac hypertrophy.     

TP53 mutations in colorectal cancer from Tunisia: relationships with site of tumor origin,microsatellite instability and KRAS mutations

Loss of TP53 function through gene mutation is a critical event in the development and progression of colorectal cancer (CRC). Here we examined 51 primary CRC tumors from Tunisia for mutations in TP53 exons 4–9 using PCR-direct sequencing. TP53 status and mutation site/type were than correlated with nuclear protein accumulation, familial and clinicopathologic variables and data on KRAS mutations and microsatellite instability (MSI-H). The TP53 mutation analysis was possible in the tumor of 47 patients and a deleterious somatic mutation has been detected in 59.6 % of the patients (28/47) including 20 (71.4 %) missense mutations, 7 nonsense mutations (25 %) and 1 (3.6 %) frameshift mutation. 89.3 % (25/28) of the detected mutations were in exons 5–8, whereas 10.7 % (3/28) were in exon 4. Among the 27 non frameshift mutations, 89 % (24/27) were transitions and 11 % (3/27) were transversions. 64.3 % (18/27) of the altered amino acids corresponded to arginine. 74 % (20/27) were G>C to A>T transitions, and more than half (14/27) occur at hotspots codons with CpG sites. TP53 mutations correlated closely with TP53 accumulation (p = 0.0090) and inversely with MSI phenotype (p = 0.0658). A KRAS somatic mutation was identified in 25 % (7/28) of the TP53 mutated tumors. All these mutations were G>A transitions in codon 12 and all the tumors with combined alterations but one were distally located and MSS. In conclusion, frequency and types of TP53 mutations and correlations with TP53 protein accumulation, and MSI were as reported for non-Tunisian patients. However, no significant associations have been detected between TP53 mutations and clinicopathological data in Tunisian patients as previously reported.