Integrative and comparative genomics analysis of early hepatocellular carcinoma differentiated from liver regeneration in young and old.

BACKGROUND: Hepatocellular carcinoma (HCC) is the third-leading cause of cancer-related deaths worldwide. It is often diagnosed at an advanced stage, and hence typically has a poor prognosis. To identify distinct molecular mechanisms for early HCC we developed a rat model of liver regeneration post-hepatectomy, as well as liver cells undergoing malignant transformation and compared them to normal liver using a microarray approach. Subsequently, we performed cross-species comparative analysis coupled with copy number alterations (CNA) of independent early human HCC microarray studies to facilitate the identification of critical regulatory modules conserved across species. RESULTS: We identified 35 signature genes conserved across species, and shared among different types of early human HCCs. Over 70% of signature genes were cancer-related, and more than 50% of the conserved genes were mapped to human genomic CNA regions. Functional annotation revealed genes already implicated in HCC, as well as novel genes which were not previously reported in liver tumors. A subset of differentially expressed genes was validated using quantitative RT-PCR. Concordance was also confirmed for a significant number of genes and pathways in five independent validation microarray datasets. Our results indicated alterations in a number of cancer related pathways, including p53, p38 MAPK, ERK/MAPK, PI3K/AKT, and TGF-beta signaling pathways, and potential critical regulatory role of MYC, ERBB2, HNF4A, and SMAD3 for early HCC transformation. CONCLUSIONS: The integrative analysis of transcriptional deregulation, genomic CNA and comparative cross species analysis brings new insights into the molecular profile of early hepatoma formation. This approach may lead to robust biomarkers for the detection of early human HCC.

Microvascular injury, thrombosis, inflammation, and apoptosis in the pathogenesis of heatstroke: a study in baboon model.

OBJECTIVE: Severe heatstroke is a leading cause of morbidity and mortality during heat waves. The pathogenesis of tissue injury, organ failure, and death in heatstroke is not well understood. METHODS AND RESULT: We investigated the pathways of heatstroke-induced tissue injury and cell death in anesthetized baboons (Papio hamadyras) subjected to environmental heat stress until core temperature attained 42.5 degrees C (moderate heatstroke; n = 3) or onset of severe heatstroke (n = 4) signaled by a fall in systolic blood pressure to < 90 mm Hg and rise in core temperature to 43.1+/-0.1 degrees C. Three sham-heated animals served as controls. Light and electron microscopy revealed widespread hemorrhage and thrombosis, transmural migration of leukocytes, and microvascular endothelium injury in severe heatstroke. Immunohistology and ultrastructural analysis demonstrated increased staining of endothelial von Willebrand factor (vWF), tissue factor (TF), and endothelial leukocyte-platelet interaction. Extensive apoptosis was noted in spleen, gut, and lung, and in hematopoeitic cells populating these organs. Double-labeling studies colocalized active caspase-3 and TF with apoptotic cells. Findings in sham-heated animals were unremarkable. CONCLUSIONS: These data suggested that microvascular injury, thrombosis, inflammation, and apoptosis may play an important role in the pathogenesis of heatstroke injury.

T313M PINK1 mutation in an extended highly consanguineous Saudi family with early-onset Parkinson disease.

BACKGROUND: To date, 5 well-confirmed genes for Parkinson disease (PD) have been identified, including 3 autosomal recessive genes: PTEN-induced putative kinase 1 (PINK1), parkin, and DJ-1. Almost nothing is known about the genetics of PD in Saudi Arabia; however, consanguineous families, not infrequent in this population, could be important in the evaluation of known PD genes and the search for new PD factors in the future. OBJECTIVE: To investigate known recessive PD genes in 5 consanguineous Saudi families with PD. DESIGN: The entire open frame as well as the untranslated region and all 5' and 3' intron-exon boundaries of the PINK1, parkin, and DJ-1 genes were sequenced in 5 probands in Saudi families. RESULTS: Four of 5 probands tested negative for PINK1, parkin, and DJ-1 mutations. However, in a large Saudi family with PD with at least 3 consanguineous marriages between first cousins, we detected a threonine to methionine substitution at codon 313 (T313M) PINK1 mutation that affected the kinase domain. Manifestations of the disease in this family included early onset (age, 28-38 years), tremulous movement, slow progression, diurnal fluctuations, bradykinesia, good response to levodopa therapy, and only mild dyskinesias. A neurologist blinded to genetic status clinically evaluated 15 family members, all older than 20 years, and diagnosed PD only in individuals who were later found to be homozygous for the T313M mutation. None of the 13 heterozygotes demonstrated any sign of PD. CONCLUSION: A homozygous T313M mutation is responsible for PD in this large Saudi family. However, the heterozygous T313M mutation does not act as a PD susceptibility factor, which is in contrast to several reports of mutations affecting only 1 PINK1 allele discovered in sporadic PD.

A proteomic analysis of excreted and circulating proteins from obese patients following two different weight-loss strategies.

Bariatric surgery is the most successful therapeutic approach to weight loss, but how it leads to weight loss, and how it resolves obesity-related complications, including type-2 diabetes, are poorly understood. This study, comprising two groups of individuals, one on a low-calorie diet (n = 5) and one undergoing bariatric surgery (n = 7), used both targeted and untargeted proteomic approaches to determine changes in protein levels pre- and post-intervention (i.e. 3-6 months later). Changes were observed in both circulating and excreted proteins following weight loss. Targeted multiplexed biochip arrays measured 12 plasma peptides/proteins involved in metabolism and inflammation: C-peptide, ferritin, interleukin-6, interleukin-1 alpha, resistin, insulin, tumor necrosis factor alpha, leptin, plasminogen-activator inhibitor-1, adiponectin, cystatin C, and C-reactive protein. Following a low-calorie diet, plasma insulin and C-reactive protein levels were significantly reduced (P = 0.045 and P = 0.030, respectively); adiponectin increased and leptin decreased following surgery (P = 0.014 and P = 0.005, respectively). Untargeted proteomic analysis employing 2D difference in-gel electrophoresis (DIGE) showed 28 protein spots with ≥1.5-fold changes in expression following weight loss by a low-calorie diet; comparison of pre- and post-intervention urine samples from the bariatric surgery group showed changes in excretion of 110 protein spots. The combination of targeted protein analysis by multiplexed arrays and an exploratory (i.e. an unbiased or discovery) proteomic assessment of hundreds of proteins offers valuable insights into the mechanistic differences between alternative weight-loss strategies. This is a powerful hypothesis-generating approach to study complex, multifactorial syndromes such as obesity. The findings that arise from these studies can then be validated in targeted, hypothesis-directed investigations.

Induction of cell proliferation in old rat liver can reset certain gene expression levels characteristic of old liver to those associated with young liver.

During the past decade, it has become increasingly clear that consistent changes in the levels of expression of a small cohort of genes accompany the aging of mammalian tissues. In many cases, these changes have been shown to generate features that are characteristic of the senescent phenotype. Previously, a small pilot study indicated that some of these changes might be reversed in rat liver, if the liver cells became malignant and were proliferating. The present study has tested the hypothesis that inducing proliferation in old rat liver can reset the levels of expression of these age-related genes to that observed in young tissue. A microarray approach was used to identify genes that exhibited the greatest changes in their expression during aging. The levels of expression of these markers were then examined in transcriptomes of both proliferating hepatomas from old animals and old rat liver lobes that had regenerated after partial hepatectomy but were again quiescent. We have found evidence that over 20 % of the aging-related genes had their levels of expression reset to young levels by stimulating proliferation, even in cells that had undergone a limited number of cell cycles and then become quiescent again. Moreover, our network analysis indicated alterations in MAPK/ERK and Jun-N-terminal kinase pathways and the potential important role of PAX3, VCAN, ARRB2, NR1H2, and ITGA5 that may provide insights into mechanisms involved in longevity and regeneration that are distinct from cancer.

Crural tunica albuginea autograft for corporoplasty: an experimental animal study of hemodynamic, histopathological, and molecular effects in the long term.

INTRODUCTION: Correction of penile deformity caused by Peyronie's disease by a variety of grafts varies in success. A long-term follow-up shows a significant number of graft scarring and erectile dysfunction. The clinical success of autologous crural tunica albuginea graft (TAG) has not resulted in wide application. AIM: To identify in healthy baboons the limitations and merits of autologous crural TAG over 1 year in a way difficult to pursue in humans. METHODS: Under general anesthesia, eight sexually active adult baboons underwent pharmacological cavernosometry (CM) and cavernosography. TAG from crus was implanted in the distal penile shaft. After 6 months, six animals were reevaluated and two were sacrificed, and the penises were excised. After 1 year, the remaining six animals were evaluated and sacrificed. The TAG and underlying corpus cavernosum (CC) were examined histologically and by Western blot analysis for nitric oxide synthase (NOS), neuronal (nNOS), endothelial (eNOS) and inducible (iNOS) isoforms, and transforming growth factor-beta1 (TGF-beta1). MAIN OUTCOME MEASURES: Sexual activity, CM, cavernosography, histopathology, and Western blot analysis. RESULTS: All animals resumed normal sexual activity 1 month postsurgery. Cavernous pressure was comparable before, at 6 months, and 1 year after surgery. A cavernovenous insufficiency developed in four animals at 6 months, and ceased in two at 1 year. Penile angulation (<20 degrees) was seen in three animals at 6 months, and an additional two at 1 year. Histologically, TAG was indistinguishable from the adjacent tunica with no fibrosis. In CC, iNOS and nNOS decreased at 1 year, whereas there was no change in TGF-beta1 levels. In TAG, there was no significant change in TGF-beta1 and eNOS levels, but there was a significant decrease in iNOS at 1 year. CONCLUSION: Autologous free TAG is associated with normal sexual activity, minimal hemodynamic changes, excellent histological outcome, and no rise in iNOS or TGF-beta1. However, cavernovenous insufficiency, mild penile angulation, and decreased nNOS persisted at 1 year.