Aberrant Distribution Research Articles

Histone Modifications and Chromatin Organization in Prostate Cancer

Epigenetic mechanisms, including histone modifications, nucleosomal remodeling and chromosomal looping, contribute to the onset and progression of prostate cancer. Recent technical advances significantly increase our understanding of the genome-wide epigenetic regulation of gene expression in prostate cancer. Aberrant genomic distribution and global level of histone modifications, nucleosome repositioning at the gene promoter and enhancer regions, as well as androgen receptor-mediated chromosomal looping may lead to the silencing of tumor suppressor genes and the activation of proto-oncogenes. In addition, androgen receptor-induced chromosomal looping facilitates recurrent gene fusion in prostate cancer. Studies in epigenetic regulation have translational implications in the identification of new biomarkers and the development of new therapies in prostate cancer.

Aberrant Distributions and Relationships Among E-cadherin, β-catenin, and Connexin 26 and 43 in Endometrioid Adenocarcinomas

During carcinogenesis, loss of intracellular cohesion is observed among cancer cells with altered expression of such adhesion molecules as E-cadherin and beta-catenin, and aberrant expression and cellular location of intercellular gap junction proteins-connexins. The aim of this study was to evaluate immunohistochemically the expression and relationship between E-cadherin and beta-catenin, and the connexins Cx26 and Cx43 in 86 endometrioid adenocarcinomas. The aberrant cytoplasmic translocation of the studied proteins was a predominant finding, whereas only a minority of cases showed normal, nuclear beta-catenin labeling or membranous distribution of the remaining molecules. E-cadherin was positively and significantly associated with beta-catenin (P=0.001, r=0.366), as was Cx26 with Cx43 (P<0.001, r=0.719), E-cadherin with Cx26 (P<0.001, r=0.413), and E-cadherin and Cx43 (P<0.001, r=0.434) in all cancers. A subgroup of endometrioid adenocarcinomas (FIGO IB+II) exclusively showed a positive significant association between the expression of beta-catenin and Cx26 (P=0.038, r=0.339). In addition, there were significantly more beta-catenin-positive carcinomas among superficially spreading cancers (FIGO IA) than among deeper invading neoplasms (FIGO IB+II) (P=0.056). The altered location of the studied proteins indicates impairment of their physiological functions. In particular, normal membranous distribution of E-cadherin and connexins is lost and replaced by abnormal cytoplasmic accumulation in most cancers, and thus intercellular ties are expected to be weakened and loosened as a consequence. In contrast, the lack of relationship between beta-catenin and connexins, E-cadherin seems to be closely associated with the expression of Cx26 and Cx43 in endometrioid adenocarcinomas.

Downregulation of Rap1GAP in Human Tumor Cells Alters Cell/Matrix and Cell/Cell Adhesion

Rap1GAP expression is decreased in human tumors. The significance of its downregulation is unknown. We show that Rap1GAP expression is decreased in primary colorectal carcinomas. To elucidate the advantages conferred on tumor cells by loss of Rap1GAP, Rap1GAP expression was silenced in human colon carcinoma cells. Suppressing Rap1GAP induced profound alterations in cell adhesion. Rap1GAP-depleted cells exhibited defects in cell/cell adhesion that included an aberrant distribution of adherens junction proteins. Depletion of Rap1GAP enhanced adhesion and spreading on collagen. Silencing of Rap expression normalized spreading and restored E-cadherin, beta-catenin, and p120-catenin to cell/cell contacts, indicating that unrestrained Rap activity underlies the alterations in cell adhesion. The defects in adherens junction protein distribution required integrin signaling as E-cadherin and p120-catenin were restored at cell/cell contacts when cells were plated on poly-l-lysine. Unexpectedly, Src activity was increased in Rap1GAP-depleted cells. Inhibition of Src impaired spreading and restored E-cadherin at cell/cell contacts. These findings provide the first evidence that Rap1GAP contributes to cell/cell adhesion and highlight a role for Rap1GAP in regulating cell/matrix and cell/cell adhesion. The frequent downregulation of Rap1GAP in epithelial tumors where alterations in cell/cell and cell/matrix adhesion are early steps in tumor dissemination supports a role for Rap1GAP depletion in tumor progression.

Redistribution of mitochondria leads to bursts of ATP production during spontaneous mouse oocyte maturation

During mammalian oocyte maturation there are marked changes in the distribution of mitochondria that supply the majority of the cellular ATP. Such redistribution of mitochondria is critical for oocyte quality, as oocytes with a poor developmental potential display aberrant mitochondrial distribution and lower ATP levels. Here we have investigated the dynamics of mitochondrial ATP production throughout spontaneous mouse oocyte maturation, using live measurements of cytosolic and mitochondrial ATP levels. We have observed three distinct increases in cytosolic ATP levels temporally associated with discrete events of oocyte maturation. These changes in cytosolic ATP levels are mirrored by changes in mitochondrial ATP levels, suggesting that mitochondrial ATP production is stimulated during oocyte maturation. Strikingly, these changes in ATP levels correlate with the distribution of mitochondria undergoing translocation to the peri-nuclear region and aggregation into clusters. Mitochondrial clustering during oocyte maturation was concomitant with the formation of long cortical microfilaments and could be disrupted by cytochalasin B treatment. Furthermore, the ATP production bursts observed during oocyte maturation were also inhibited by cytochalasin B suggesting that mitochondrial ATP production is stimulated during oocyte maturation by microfilament-driven, sub-cellular targeting of mitochondria. J. Cell. Physiol. 224: 672–680, 2010. © 2010 Wiley-Liss, Inc.

Premature expression of KCC2 in embryonic mice perturbs neural development by an ion transport‐independent mechanism

During neuronal maturation, the neuron-specific K-Cl co-transporter KCC2 lowers the intracellular chloride and thereby renders GABAergic transmission hyperpolarizing. Independently of its role as a co-transporter, KCC2 plays a crucial role in the maturation of dendritic spines, most probably via an interaction with the cytoskeleton-associated protein 4.1N. In this study, we show that neural-specific overexpression of KCC2 impairs the development of the neural tube- and neural crest-related structures in mouse embryos. At early stages (E9.5-11.5), the transgenic embryos had a thinner neural tube and abnormal body curvature. They displayed a reduced neuronal differentiation and altered neural crest cell pattern. At later stages (E11.5-15.5), the transgenic embryos had smaller brain structures and a distinctive cleft palate. Similar results were obtained using overexpression of a transport-inactive N-terminal-deleted variant of KCC2, implying that the effects were not dependent on KCC2's role as a K-Cl co-transporter. Interestingly, the neural tube of transgenic embryos had an aberrant cytoplasmic distribution of 4.1N and actin. This was corroborated in a neural stem cell line with ectopic expression of KCC2. Embryo phenotype and cell morphology were unaffected by a mutated variant of KCC2 which is unable to bind 4.1N. These results point to a role of KCC2 in neuronal differentiation and migration during early development mediated by its direct structural interactions with the neuronal cytoskeleton.

Abstract 3466: Increased steady-state levels of microtubules in aberrant distributions in Ni+2 ion-/MCA-transformed C3H/10T1/2 mouse embryo cell lines

Abstract We previously found amplification of the Ect-2 proto-oncogene, and higher steady-state levels of ect-2 mRNA and protein, in Ni+2 ion- and MCA-transformed C3H/10T1/2 (10T1/2) mouse embryo fibroblast cell lines. Here, we tested the hypotheses that there were 1) increased steady-state levels of microtubules in Ni+2/MCA-transformed 10T1/2 cell lines, 2) aberrant distributions of microtubules in transformed cell lines, and 3) increases in cross sectional areas of transformed cell lines. Log phase Ni+2 ion- and MCA-transformed and non-transformed 10T1/2 mouse embryo cells were stained separately with fluorescent antibodies to alpha tubulin or beta tubulin, then visualized using confocal microscopy (n=8). The number of microtubules/cell and cross sectional areas of stained cells were quantified using the volocity image analysis program, and ratios of (microtubules in transformed cell lines/microtubules in non-transformed 10T1/2 cells) were determined. In non-transformed 10T1/2 cells, microtubules were distributed homogeneously. In transformed cells, microtubules were often present as large aggregates in cells, and shapes of transformed cells were aberrant compared to shapes of non-transformed 10T1/2 cells. Compared to non-transformed 10T1/2 cells, MCA Cl 16, NiS3A1, NiS7A1 and NiO2C3 cell lines had a 2.1 (p &amp;lt; 0.001), 1.5 (p &amp;gt; 0.05), 1.8 (p &amp;gt; 0.05) and 1.9 (p &amp;lt; 0.05) fold higher fluorescent intensity for alpha tubulin, respectively. Compared to non-transformed 10T1/2 cells, MCA Cl 16, NiS3A1, NiS7A1 and NiO2C3 had 2.5 (p &amp;lt; 0.05) 1.7 (p&amp;lt;0.05), 1.8 (p&amp;gt;0.05) and 1.1 (p&amp;lt;0.05) fold-higher beta tubulin fluorescence intensity relative to 10T1/2 cells. Transformed cell lines showed 1.9 - 2.5 significant (p&amp;lt;0.1) fold-increases in cross-sectional area compared to non-transformed 10T1/2 cells (n = 8). We concluded MCA-/Ni+2 ion-transformed 10T1/2 cells had higher steady-sate levels of microtubules, present in aberrant distributions, and transformed cells were larger than non-transformed cells. Supported by Grant ES03341 from U. S. N. I. H., funding from the M. S. Program in the Dept. of Molecular Microbiology/Immunology at USC, and discretionary funding, to JRL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3466.

Sperm equatorial segment protein 1, SPESP1, is required for fully fertile sperm in mouse

Mammalian fertilization is a multistep process that culminates in the fusion of the sperm and egg plasma membrane. It is widely accepted that the equatorial segment of the acrosome-reacted sperm is important in initiating fusion with the egg plasma membrane during fertilization. There are various proteins known to be distributed only in the equatorial segment of sperm. The role of these proteins must be clarified to understand the membrane fusion process. We produced a mouse line that lacked SPESP1 (sperm equatorial segment protein 1) and analyzed the fertilizing ability of the sperm. The average number of pups that were fathered by Spesp1(+/-) and Spesp1(-/-) males was significantly lower than that of wild-type fathers. In these mouse lines, fewer sperm were found to migrate into oviducts and fewer eggs were fertilized. The Spesp1(+/-) and Spesp1(-/-) sperm showed a lower fusing ability compared with the wild-type sperm. The disruption of Spesp1 was shown to cause an aberrant distribution of various sperm proteins. Moreover, scanning electron microscopy revealed that the membrane in the equatorial segment area, which usually forms an acrosomal sheath, disappears after acrosome reaction in Spesp1-deficient mice. It was demonstrated that SPESP1 is necessary to produce the fully 'fusion competent' sperm.

DNA Demethylating Antineoplastic Strategies: A Comparative Point of View

Despite the involvement of genetic alterations in neoplastic cell transformation, it is increasingly evident that abnormal epigenetic patterns, such as those affecting DNA methylation and histone posttranslational modifications (PTMs), play an essential role in the early stages of tumor development. This finding, together with the evidence that epigenetic changes are reversible, enabled the development of new antineoplastic therapeutic approaches known as epigenetic therapies. Epigenetic modifications are involved in the control of gene expression, and their aberrant distribution is thought to participate in neoplastic transformation by causing the deregulation of crucial cellular pathways. Epigenetic drugs are able to revert the defective gene expression profile of cancer cells and, consequently, reestablish normal molecular pathways. Considering the emerging interest in epigenetic therapeutics, this review focuses on the approaches affecting DNA methylation, evaluates novel strategies and those already approved for clinical use, and compares their therapeutic potential.

The Effect of Insulin Signaling on Female Reproductive Function Independent of Adiposity and Hyperglycemia

Physiological states of insulin resistance such as obesity and diabetes have been linked to abnormalities in female reproductive function. However, it is difficult to distinguish the direct effects of impaired insulin signaling from those of adiposity or hyperglycemia because these conditions often coexist in human syndromes and animal models of insulin resistance. In this study, we used lean, normoglycemic mouse lines with differing degrees of hyperinsulinemia and insulin receptor (Insr) expression to dissect the effects of altered insulin signaling on female reproduction. All three mouse lines [Ttr-Insr(-/-), Insr(+/-), and Insr(+/+) (wild type)] are able to maintain fertility. However, the insulin-resistant and hyperinsulinemic mice demonstrate altered duration of estrous cycles as well as aberrant distribution and morphology of ovarian follicles. These effects appear to be independent of hyperandrogenism in the mice. Pregnancy studies indicate decreased success in early progression of gestation. In successful pregnancies, decreased embryo weights and increased placental calcification also implicate altered insulin signaling in later gestational effects. Thus, abnormal insulin signaling, independent of adipose tissue mass, adipokine expression levels, and hyperglycemia, can affect parameters of the female hypothalamic-pituitary-gonadal axis and pregnancy outcomes.

Loss of SFRP1 Expression is Associated with Aberrant β-Catenin Distribution and Tumor Progression in Mucoepidermoid Carcinoma of Salivary Glands

Cytoplasmic and nuclear accumulation of beta-catenin in mucoepidermoid carcinoma (MEC) is frequently noted, but the mechanism is unknown. The methylation status of adenomatous polyposis coli (APC) and secreted frizzled-related proteins (SFRPs) was examined by methylation-specific polymerase chain reaction (MSP) assay. The association of SFRP1, beta-catenin, and cyclin D1 expression in MEC was evaluated by immunohistochemical staining. A high percentage of methylation in APC and the SFRP genes was found in MEC compared with adjacent normal tissues, in which SFRP1 (58.6%) was the most frequent methylated gene. Moreover, abundant expression of SFRP1 was noted in normal tissues, whereas reduced SFRP1 expression was detected in 71.7% (33/46) of MECs. There was significant association between methylation and reduced expression of SFRP1. Cytoplasmic/nuclear (C/N) beta-catenin and high cyclin D1 expression were found in 13/55 (23.6%) and 36/55 (65.5%) of cases, respectively. There was significant correlation between C/N beta-catenin expression and reduced SFRP1 expression (P = 0.009). In addition, SFRP1 and beta-catenin expression correlated with tumor malignancy index such as tumor grade and stage. Overall patient survival was significantly worse in patients with reduced SFRP1 and C/N beta-catenin expression (P = 0.009 and P = 0.002, respectively). Methylation of the SFRP1 gene was the major cause of reduced SFRP1 expression. Reduced SFRP1 led to C/N accumulation of beta-catenin and was associated with tumor malignancy. Therefore, examination of SFRP1 expression and beta-catenin location could be useful predictors of tumor progression and prognosis in patients with MEC.

Nuclear or cytoplasmic localization of Bag-1 distinctly correlates with pathologic behavior and outcome of gastric carcinomas

Bag-1 is an antiapoptotic protein with its altered expression and localization in malignancies. To clarify the role of Bag-1 in gastric carcinogenesis, its expression was examined by immunohistochemistry and in situ hybridization on a tissue microarray containing gastric carcinomas, adjacent nonneoplastic mucosa (NNM), adenomas, intestinal metaplasia (IM), or gastritis. Gastric carcinoma tissue and cell lines were studied for Bag-1 expression by Western blot and reverse transcriptase-polymerase chain reaction (RT-PCR). The results demonstrated that Bag-1 proteins were differentially expressed in the nucleus or cytosol of MKN28, AGS, MKN45, KATO-III, or HGC-27 cell lines, despite similar levels of messenger RNA (mRNA) expression. The Bag-1 mRNA overexpression was detectable in 73.3% of 15 gastric carcinomas without significant difference in its encoding products' levels. The nuclear Bag-1 expression gradually decreased from gastritis, IM, adenoma to carcinoma (P < .05), and negatively correlated with lymphatic invasion or lymph node metastasis, cytoplasmic Bag-1 expression, negative parafibromin expression, and poor prognosis (P < .05). Cytoplasmic Bag-1 was weakly immunoreactive in carcinomas, compared with gastritis (P < .05), and positively associated with invasive depth and poor prognosis of the carcinoma (P < .05). The positive rate of Bag-1 mRNA expression was higher in adjacent IMs than carcinomas or adjacent NNM (P < .05). Bag-1 mRNA was expressed more in carcinomas from female patients than the male counterparts (P < .05). There was a positive correlation of Bag-1 mRNA expression with invasive depth and venous invasion (P < .05). Our study indicated that aberrant expression and subcellular distribution of Bag-1 might play an important role in the malignant transformation of gastric epithelial cells and should be considered as a biomarker for gastric carcinogenesis, subsequent progression, and prognosis.

Secreted semaphorins control spine distribution and morphogenesis in the postnatal CNS

The majority of excitatory synapses in the mammalian CNS are formed on dendritic spines1, and spine morphology and distribution are critical for synaptic transmission2–6, synaptic integration and plasticity7. Here, we show that a secreted semaphorin, Sema3F, is a negative regulator of spine development and synaptic structure. Mice with null mutations in genes encoding Sema3F, and its holoreceptor components neuropilin-2 (Npn-2) and plexinA3 (PlexA3), exhibit increased dentate gyrus (DG) granule cell (GC) and cortical layer V pyramidal neuron spine number and size, and also aberrant spine distribution. Moreover, Sema3F promotes loss of spines and excitatory synapses in dissociated neurons in vitro, and in Npn-2−/− brain slices cortical layer V and DG GCs exhibit increased mEPSC frequency. In contrast, a distinct Sema3A–Npn-1/PlexA4 signaling cascade controls basal dendritic arborization in layer V cortical neurons but does not influence spine morphogenesis or distribution. These disparate effects of secreted semaphorins are reflected in the restricted dendritic localization of Npn-2 to apical dendrites and of Npn-1 to all dendrites of cortical pyramidal neurons. Therefore, Sema3F signaling controls spine distribution along select dendritic processes, and distinct secreted semaphorin signaling events orchestrate CNS connectivity through the differential control of spine morphogenesis, synapse formation, and the elaboration of dendritic morphology.

Oncocytic versus mitochondrion‐rich follicular thyroid tumours: should we make a difference?

To separate true oncocytic neoplasms from mitochondrion-rich non-oncocytic lesions based on the intracellular relationship between major cell organelles, and to establish the diagnostic and clinical relevance of this distinction. Tissue samples from 276 follicular adenomas, 194 follicular carcinomas, 162 normal thyroids and 296 non-neoplastic lesions were classified as conventional, mitochondrion-rich or oncocytic based on the immunohistochemically assessed quantity and intracellular distribution of mitochondria and endoplasmic reticulum (ER) and nuclear position. Pathological and clinical features were compared among the groups. In oncocytes, densely packed mitochondria resulted in homogeneous immunolabelling of basal cytoplasmic regions, whereas ER and the nuclei were typically displaced to the apical position. This aberrant organelle distribution was not observed in non-oncocytes, which allowed reliable distinction between oncocytic and mitochondrion-rich lesions. Clinically, mitochondrial increase in non-oncocytic lesions was associated with neoplasia, malignancy and higher cancer recurrence rates. Similar correlation, albeit less pronounced, was observed within the oncocytic tumour group. By contrast, oncocytic change per se was not associated with neoplasia, malignancy or cancer aggressiveness. True oncocytic neoplasms can be distinguished from mitochondrion-rich non-oncocytic tumours based on aberrant distribution of all major cell organelles. This distinction has immediate clinical relevance and should be implemented in practice.

Aberrant distribution of junctional complex components in retinoic acid receptor alpha‐deficient mice

Retinoic acid receptor alpha (RARalpha)-deficient mice are sterile, with abnormalities in the progression of spermatogenesis and spermiogenesis. In this study, we investigated whether defective retinoid signaling involved at least in part, disrupted cell-cell interactions. Hypertonic fixation approaches revealed defects in the integrity of the Sertoli-cell barrier in the tubules of RARalpha-deficient testes. Dye transfer experiments further revealed that coupling between cells from the basal to adluminal compartments was aberrant. There were also differences in the expression of several known retinoic acid (RA)-responsive genes encoding structural components of tight junctions and gap junctions. Immunostaining demonstrated a delay in the incorporation of zonula occludens (ZO-1), a peripheral component protein of tight junctions, into the Sertoli cell tight junctions. Markedly reduced expression of connexin-40 in mutant pachytene spermatocytes and round spermatids was found by in situ hybridization. An ectopic distribution of vimentin and disrupted cyclic expression of vimentin, which is usually tightly regulated during spermiogenesis, was found in RARalpha-deficient testes at all ages examined. Thus, the specific defects in spermiogenesis in RARalpha-deficient testes may correlate with a disrupted cyclic expression of RA-responsive structural components, including vimentin, a downregulation of connexin-40 in spermatogenic cells, and delayed assembly of ZO-1 into Sertoli cell tight junctions. Interestingly, bioinformatic analysis revealed that many genes that are components of tight junctions and gap junctions contained potential retinoic acid response element binding sites.

Hamartin Variants That Are Frequent in Focal Dysplasias and Cortical Tubers Have Reduced Tuberin Binding and Aberrant Subcellular Distribution In Vitro

Focal cortical dysplasia type IIb is characterized by epilepsy-associated malformations that are often composed of balloon cells and dysplastic neurons. There are many histopathologic similarities between focal cortical dysplasia type IIb and cortical tubers in tuberous sclerosis complex (TSC), an autosomal-dominant phakomatosis caused by mutations in the TSC1 or TSC2 genes that encode hamartin and tuberin. We previously found that an allelic variant of TSC1 (hamartin) is increased in focal cortical dysplasia type IIb. Here, we investigated the subcellular localization of hamartin and its interaction with tuberin in vitro. Coimmunoprecipitation assays with tuberin revealed reduced tuberin binding of hamartin compared with wild-type hamartin. Tuberin binding was also reduced for 2 TSC1 stop mutants (hamartin and hamartin) that are present in brain lesions of TSC patients. Colocalization assays of hamartin and tuberin were performed in HEK293T cells, and the subcellular localization of the hamartin variants were studied using immunocytochemistry. There was an impairment of tuberin binding of hamartin and aberrant nuclear distribution of hamartin in these cells, whereas hamartin and hamartin were, like wild-type tuberin, localized in the cytoplasm. These data suggest a fundamental functional impairment of hamartin and the 2 TSC1 stop mutants hamartin and hamartin in vitro. Future studies will be needed to characterize the roles of these TSC1 sequence variants in the genesis of dysplastic epileptogenic developmental brain lesions.

Decreased PPARγ Expression Compromises Perigonadal-Specific Fat Deposition and Insulin Sensitivity

Mutations and polymorphisms in PPARG have been linked to adiposity and partial lipodystrophy in humans. However, how disturbances in PPARG lead to depot-specific effects on adipose tissue, as shown by the characteristic aberrant fat distribution in patients, remains unclear. By manipulating the 3′-untranslated region of the Pparg gene, we have generated mice with peroxisome proliferator-activated receptor γ (PPARγ) gene expression ranging from 25% to 100% normal. Basal levels of PPARγ transcripts between 50% and approximately 100% had no significant effect on body weight, fat mass, and insulin sensitivity. In contrast, mice with 25% normal PPARγ expression exhibited reduced body weight and total fat mass, insulin resistance, and dyslipidemia. Interestingly, fat mass was selectively reduced in perigonadal depot without significant changes in inguinal and other depots. Expression of adipogenic factor CCAAT enhancer binding protein-α and some other metabolic genes containing peroxisome proliferator response element were reduced in a perigonadal depot-specific fashion. This was further associated with depot-specific reduction in the expression of adipokines, increased expression of TNFα, and increased ectopic lipid deposition in muscles. Together, these results underscore the differential sensitivity of the individual fat depots on PPARγ availability as an underlying mechanism of partial lipodystrophy.

Axonal ligation induces transient redistribution of TDP-43 in brainstem motor neurons

Nuclear exclusion of TAR DNA binding protein 43 (TDP-43) and formation of cytosolic aggregates are a pathological characteristic of amyotrophic lateral sclerosis (ALS). However, the molecular basis of the aberrant distribution of TDP-43 remains elusive. Here, we show evidence that axonal ligation induced transient nuclear exclusion and peripheral accumulation of TDP-43, without apparent cytosolic aggregates in hypoglossal neurons in mice. Immunohistochemistry showed marked loss of nuclear TDP-43 7–14 days after ligation, which was accompanied by reduction of choline acetyltransferase (ChAT). TDP-43 staining was restored in the nucleus on day 28 exclusively in the neurons with normalized ChAT expression. We also showed that importin β, which was shown to mediate nuclear transport of TDP-43 was downregulated transiently by nerve ligation. The analysis of the peripheral nerves proximal to the ligation revealed that TDP-43 markedly accumulated with a concomitant decrease in active autophagosome. Moreover, we showed that TDP-43 was present in the microsome fraction containing endoplasmic reticulum (ER) or autophagosomes in the brainstem section, indicating that TDP-43 is axonally transported with vesicles. These results indicate that axonal damage is associated with redistribution of TDP-43 through the combination of defective axonal autophagy periphery and the impaired nuclear transport system in the soma. Moreover, it was also shown that transient redistribution of TDP-43 does not prevent motor neurons from axonal regeneration. Therefore, our data suggest that the subcellular distribution of TDP-43 correlates to the innervation status of motor neurons, which may be governed by unidentified cause of ALS.

Diminished dosage of 22q11 genes disrupts neurogenesis and cortical development in a mouse model of 22q11 deletion/DiGeorge syndrome

The 22q11 deletion (or DiGeorge) syndrome (22q11DS), the result of a 1.5- to 3-megabase hemizygous deletion on human chromosome 22, results in dramatically increased susceptibility for "diseases of cortical connectivity" thought to arise during development, including schizophrenia and autism. We show that diminished dosage of the genes deleted in the 1.5-megabase 22q11 minimal critical deleted region in a mouse model of 22q11DS specifically compromises neurogenesis and subsequent differentiation in the cerebral cortex. Proliferation of basal, but not apical, progenitors is disrupted, and subsequently, the frequency of layer 2/3, but not layer 5/6, projection neurons is altered. This change is paralleled by aberrant distribution of parvalbumin-labeled interneurons in upper and lower cortical layers. Deletion of Tbx1 or Prodh (22q11 genes independently associated with 22q11DS phenotypes) does not similarly disrupt basal progenitors. However, expression analysis implicates additional 22q11 genes that are selectively expressed in cortical precursors. Thus, diminished 22q11 gene dosage disrupts cortical neurogenesis and interneuron migration. Such developmental disruption may alter cortical circuitry and establish vulnerability for developmental disorders, including schizophrenia and autism.

Decreased PPARγ Expression Compromises Perigonadal-Specific Fat Deposition and Insulin Sensitivity

Mutations and polymorphisms in PPARG have been linked to adiposity and partial lipodystrophy in humans. However, how disturbances in PPARG lead to depot-specific effects on adipose tissue, as shown by the characteristic aberrant fat distribution in patients, remains unclear. By manipulating the 3'-untranslated region of the Pparg gene, we have generated mice with peroxisome proliferator-activated receptor gamma (PPAR gamma) gene expression ranging from 25% to 100% normal. Basal levels of PPAR gamma transcripts between 50% and approximately 100% had no significant effect on body weight, fat mass, and insulin sensitivity. In contrast, mice with 25% normal PPAR gamma expression exhibited reduced body weight and total fat mass, insulin resistance, and dyslipidemia. Interestingly, fat mass was selectively reduced in perigonadal depot without significant changes in inguinal and other depots. Expression of adipogenic factor CCAAT enhancer binding protein-alpha and some other metabolic genes containing peroxisome proliferator response element were reduced in a perigonadal depot-specific fashion. This was further associated with depot-specific reduction in the expression of adipokines, increased expression of TNFalpha, and increased ectopic lipid deposition in muscles. Together, these results underscore the differential sensitivity of the individual fat depots on PPAR gamma availability as an underlying mechanism of partial lipodystrophy.

Cell lines derived from human parthenogenetic embryos can display aberrant centriole distribution and altered expression levels of mitotic spindle check-point transcripts.

Human parthenogenetic embryos have recently been proposed as an alternative, less controversial source of embryonic stem cell (ESC) lines; however many aspects related to the biology of parthenogenetic embryos and parthenogenetic derived cell lines still need to be elucidated. We present here results on human cell lines (HP1 and HP3) derived from blastocysts obtained by oocyte parthenogenetic activation. Cell lines showed typical ESC morphology, expressed Oct-4, Nanog, Sox-2, Rex-1, alkaline phosphatase, SSEA-4, TRA 1-81 and had high telomerase activity. Expression of genes specific for different embryonic germ layers was detected from HP cells differentiated upon embryoid body (EBs) formation. Furthermore, when cultured in appropriate conditions, HP cell lines were able to differentiate into mature cell types of the neural and hematopoietic lineages. However, the injection of undifferentiated HP cells in immunodeficient mice resulted either in poor differentiation or in tumour formation with the morphological characteristics of myofibrosarcomas. Further analysis of HP cells indicated aberrant levels of molecules related to spindle formation as well as the presence of an abnormal number of centrioles and autophagic activity. Our results confirm and extend the notion that human parthenogenetic stem cells can be derived and can differentiate in mature cell types, but also highlight the possibility that, alteration of the proliferation mechanisms may occur in these cells, suggesting great caution if a therapeutic use of this kind of stem cells is considered.