Lower Level Of Cell Proliferation Research Articles

Chemopreventive effect of pomegranate and cocoa extracts on ultraviolet radiation-induced photocarcinogenesis in SKH-1 mice.

Non-melanoma skin cancer (NMSC) has a high and increasing incidence all over the world. Solar radiation is the main aetiology for humans. Although most research into photocarcinogenesis uses UVB as a source of radiation, UVA is also carcinogenic in long term. Pomegranate (PGE) and cocoa (CE) extracts have been used for medicinal purposes for time immemorial. Recently, it has been claimed that some of their properties may be an effective preventative measure against photocarcinogenesis and photoaging, but to date in vivo models have not been tested using RUVA, the objective of the present work. A lower incidence of lesions was observed in SKH-1 mice treated with PGE (p<0.001), and lower incidence of invasive squamous carcinoma in both treatment groups (p<0.001 for PGE and p<0.05 for CE); the PGE group also showed a lower level of cell proliferation than the control group (p<0.001). Significantly greater p53 alteration was observed in the control group than the treatment groups (p<0.001 for PGE and p = 0.05 for CE). No significant differences were found in relation to TIMP-1 and MMP-9. Taken together, the results suggest that oral feeding of PGE and CE to SKH-1 mice affords substantial protection against the adverse effects of RUVA, especially PGE.

Differences in Cell Proliferation and Craniofacial Phenotype of Closely Related Species in the Pupfish Genus Cyprinodon.

Understanding the genetic basis for phenotypic differences is fundamental to the study of macroevolutionary patterns of biological diversity. While technological advances in DNA sequencing have made researching genetic variation in wild taxa routine, fully understanding how these variants affect phenotype requires taking the next step to investigate how genetic changes alter cell and tissue interactions that ultimately produce phenotypes. In this article, we investigate a role for cell proliferation as a developmental source of craniofacial diversity in a radiation of 3 species of Cyprinodon from San Salvador Island, Bahamas. Patterns of cell proliferation in the heads of hatching-age fish differ among species of Cyprinodon, and correlate with differences in allometric growth rate among the jaws of 3 distinct species. Regional patterns of cell proliferation in the head are complex, resulting in an unintuitive result in which lower levels of cell proliferation in the posterior head region are associated with the development of relatively larger jaws in one species. We combine these data with previously published morphological and genomic data to show how studying the mechanisms generating phenotype at the cellular and tissue levels of biological organization can help mechanistically link genomic studies with classic morphological studies.

Abstract 1574: Circadian clock gene expression and lethal prostate cancer outcomes

Abstract Background: Converging evidence points to a potential role of disruption of the circadian rhythm in prostate cancer progression. There is limited human data in prostate tissue examining the biological consequences of alterations in genes involved in circadian rhythm on outcomes among prostate cancer patients. Methods: To test the hypothesis that tissue expression of circadian-related genes was associated with prostate cancer outcomes, we leveraged data from the Physicians' Health Study (PHS) and the Health Professionals Follow-Up Study (HPFS) prostate tumor cohorts. Whole transcriptome expression profiling was performed on 404 prostate cancer cases, including 113 lethal cases (metastatic disease or prostate cancer death) and 291 indolent cases (&amp;gt;8 years from diagnosis without evidence of metastasis). Using logistic regression models, we assessed whether mRNA expression levels in tumor (N=404) or paired normal prostate tissue (N=202) of eleven circadian-related genes (AANAT, CLOCK, CRY1, CRY2, CSNK1E, MTNR1B, NPAS2, OPN4, PER1, PER2, PER3) were associated with lethal prostate cancer, stage (T2 vs. T3+), Gleason grade at diagnosis (&amp;lt;8 vs. ≥8), and biomarkers of angiogenesis and apoptosis. We calculated odds ratios and 95% confidence intervals (95% CIs) per standard deviation increase in gene expression. Results: Pathway analyses showed a statistically significant association between the eleven circadian genes and lethal disease (global test p-value = 5.1e-05). On an individual gene level, men with higher tumor expression of Period Circadian Regulator 1 (PER1) gene had a reduced risk of lethal disease, independent of grade and stage (OR: 0.75, 95% CI: 0.59-0.97). High tumor expression of PER1 (OR: 0.72, 95% CI: 0.57-0.91) and Cryptochrome Circadian Regulator 2 (CRY2) (OR: 0.79, 95% CI: 0.63-1.00) were associated with lower Gleason grade tumors. High tumor expression of PER1 was associated with lower levels of cell proliferation and lower levels of angiogenesis markers; Neuronal PAS Domain Protein 2 (NPAS2) was associated with markers of apoptosis. For none of the genes was expression in normal prostate tissue associated with lethal prostate cancer (global test p-value = 0.06). Discussion: We found higher expression of several of the circadian related genes were associated with less aggressive prostate cancer features. In line with these findings, PER1 has been suggested to be a tumor suppressor in previous studies. This supports the idea that maintenance of circadian clock function may protect tumor progression. Citation Format: Sarah C. Markt, Ericka Ebot, Iona Cheng, Lynne Wilkens, Ayesha Shafi, Karen Knudsen, Kathryn Penney, Lorelei Mucci, Travis Gerke. Circadian clock gene expression and lethal prostate cancer outcomes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1574.

Histone Demethylase JMJD2D Interacts With β-Catenin to Induce Transcription and Activate Colorectal Cancer Cell Proliferation and Tumor Growth in Mice

Wnt signaling contributes to the development of colorectal cancer (CRC). We studied interactions between lysine demethylase 4D (KDM4D or JMJD2D) and β-catenin, a mediator of Wnt signaling, in CRC cell lines and the effects on tumor formation in mice. We obtained colorectal tumor specimens and surrounding nontumor colon tissues (controls) from patients undergoing surgery in China; levels of JMJD2D were measured by immunohistochemical or immunoblot analysis. JMJD2D expression was knocked down in CRC (CT26, HCT116, and SW480 cells) using small hairpin RNAs, and cells were analyzed with viability, flow cytometry, colony formation, andtranswell migration and invasion assays. Cells were alsogrown as tumor xenografts in nude mice or injected into tail veins or spleens of mice, and metastases were measured. We performed promoter activity, co-immunoprecipitation, and chromatin immunoprecipitation assays. We also performed studies with Apcmin/+ and JMJD2D-knockout mice; these mice were crossed, and colorectal tumor formation in offspring (Apcmin/+Jmjd2d+/+ and Apcmin/+Jmjd2d-/-) was analyzed. JMJD2D-knockout and wild-type (control) mice were given azoxymethane followed by dextran sodium sulfateto induce colitis-associated CRC; some mice were given the JMJD2D inhibitor 5-chloro-8-hydroxyquinoline (5-c-8HQ) or vehicle to examine the effects of 5-c-8HQ on intestinal tumor formation. Levels of JMJD2D were significantly higher in human colorectal tumors than in control tissues and correlated with levels of proliferating cell nuclear antigen. JMJD2D knockdown reduced CRC cell proliferation, migration, and invasion, as well as growth of xenograft tumors and formation of metastases in mice. JMJD2D was required for expression of β-catenin in CRC cell lines; ectopic expression of JMJD2D increased the promoter activities of genes regulated by β-catenin (MYC, CCND1, MMP2, and MMP9). We found that JMJD2D and β-catenin interacted physically and that JMJD2D demethylated H3K9me3 at promoters of β-catenin target genes. JMJD2D-knockout micedeveloped fewer colitis-associated colorectal tumors than control mice, and their tumor tissues had lower levels of β-catenin, MYC, cyclin D1, and proliferating cell nuclear antigen than tumorsfrom control mice. Apcmin/+Jmjd2d-/- mice developed fewer and smaller colon tumors than Apcmin/+ mice. Mice given 5-c-8HQ developed smaller and fewer colitis-associated tumors, with lower levels of cell proliferation, than mice given vehicle. Apcmin/+ mice given 5-c-8HQ also developed fewer tumors in intestines and colons than mice given vehicle. Levels of the histone demethylase JMJD2D are increased in human colorectal tumors compared with nontumor colon tissues. JMJD2D interacts with β-catenin to activate transcription of its target genes and promote CRC cell proliferation, migration, and invasion, as well as formation of colorectal tumors in mice.

Limb patterning genes and heterochronic development of the emu wing bud.

BackgroundThe forelimb of the flightless emu is a vestigial structure, with greatly reduced wing elements and digit loss. To explore the molecular and cellular mechanisms associated with the evolution of vestigial wings and loss of flight in the emu, key limb patterning genes were examined in developing embryos.MethodsLimb development was compared in emu versus chicken embryos. Immunostaining for cell proliferation markers was used to analyze growth of the emu forelimb and hindlimb buds. Expression patterns of limb patterning genes were studied, using whole-mount in situ hybridization (for mRNA localization) and RNA-seq (for mRNA expression levels).ResultsThe forelimb of the emu embryo showed heterochronic development compared to that in the chicken, with the forelimb bud being retarded in its development. Early outgrowth of the emu forelimb bud is characterized by a lower level of cell proliferation compared the hindlimb bud, as assessed by PH3 immunostaining. In contrast, there were no obvious differences in apoptosis in forelimb versus hindlimb buds (cleaved caspase 3 staining). Most key patterning genes were expressed in emu forelimb buds similarly to that observed in the chicken, but with smaller expression domains. However, expression of Sonic Hedgehog (Shh) mRNA, which is central to anterior–posterior axis development, was delayed in the emu forelimb bud relative to other patterning genes. Regulators of Shh expression, Gli3 and HoxD13, also showed altered expression levels in the emu forelimb bud.ConclusionsThese data reveal heterochronic but otherwise normal expression of most patterning genes in the emu vestigial forelimb. Delayed Shh expression may be related to the small and vestigial structure of the emu forelimb bud. However, the genetic mechanism driving retarded emu wing development is likely to rest within the forelimb field of the lateral plate mesoderm, predating the expression of patterning genes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13227-016-0063-5) contains supplementary material, which is available to authorized users.

Survival of motor neurone protein is required for normal postnatal development of the spleen.

Spinal muscular atrophy (SMA), traditionally described as a predominantly childhood form of motor neurone disease, is the leading genetic cause of infant mortality. Although motor neurones are undoubtedly the primary affected cell type, the severe infantile form of SMA (Type I SMA) is now widely recognised to represent a multisystem disorder where a variety of organs and systems in the body are also affected. Here, we report that the spleen is disproportionately small in the 'Taiwanese' murine model of severe SMA (Smn-/- ;SMN2tg/0 ), correlated to low levels of cell proliferation and increased cell death. Spleen lacks its distinctive red appearance and presents with a degenerated capsule and a disorganised fibrotic architecture. Histologically distinct white pulp failed to form and this was reflected in an almost complete absence of B lymphocytes necessary for normal immune function. In addition, megakaryoctyes persisted in the red pulp. However, the vascular density remained unchanged in SMA spleen. Assessment of the spleen in SMA patients with the infantile form of the disease indicated a range of pathologies. We conclude that development of the spleen fails to occur normally in SMA mouse models and human patients. Thus, further analysis of immune function is likely to be required to fully understand the full extent of systemic disease pathology in SMA.

P75 neurotrophin receptor regulates differential mineralization of rat ectomesenchymal stem cells.

The aim of this study was to investigate whether p75NTR (p75 neurotrophin receptor) regulates differential mineralization capacity of rEMSCs (rat ectomesenchymal stem cells) and underlying mechanisms associated with Mage-D1 (melanoma-associated antigens-D1). Immunohistochemical staining of p75NTR in developing tooth germs was performed on E12.5d (embryonic 12.5days) and E19.5d (embryonic 19.5days). E12.5d EMSCs and E19.5d EMSCs were isolated in the same pregnant Sprague-Dawley rats from embryonic maxillofacial processes and tooth germs. p75NTR small-interfering RNA, p75NTR overexpression plasmid, Mage-D1 small-interfering RNA and recombined rat NGF were used to transfect cells. p75NTR was expressed in epithelial-mesenchymal interaction areas at E12.5d and E19.5d tooth germ development stages. E19.5d EMSCs had higher p75NTR expression levels and differential mineralization capacity but lower levels of cell proliferation. Under induction by mineralized culture medium, the potential of differential mineralization had identical trends in regulation of p75NTR in EMSCs; Mage-D1 did not fluctuate and TrkA was not expressed. Binding of p75NTR and Mage-D1 were detected. Mage-D1 knockdown significantly down-regulated expression of related genes, which NGF could not rescue. p75NTR participated in tooth germ development stages and mediated differential mineralization of EMSCs. p75NTR played a critical role in regulating the potential of differential mineralization of EMSCs. Mage-D1 seemed to act as a bridge in the underlying mechanism of effects of p75NTR.

Species differences in behavior and cell proliferation/survival in the adult brains of female meadow and prairie voles

Microtine rodents display diverse patterns of social organization and behaviors, and thus provide a useful model for studying the effects of the social environment on physiology and behavior. The current study compared the species differences and the effects of oxytocin (OT) on anxiety-like, social affiliation, and social recognition behaviors in female meadow voles (Microtus pennsylvanicus) and prairie voles (Microtus ochrogaster). Furthermore, cell proliferation and survival in the brains of adult female meadow and prairie voles were compared. We found that female meadow voles displayed a higher level of anxiety-like behavior but lower levels of social affiliation and social recognition compared to female prairie voles. In addition, meadow voles showed lower levels of cell proliferation (measured by Ki67 staining) and cell survival (measured by BrdU staining) in the ventromedial hypothalamus (VMH) and amygdala (AMY), but not the dentate gyrus of the hippocampus (DG), than prairie voles. Interestingly, the numbers of new cells in the VMH and AMY, but not DG, also correlated with anxiety-like, social affiliation, and social recognition behaviors in a brain region-specific manner. Finally, central OT treatment (200ng/kg, icv) did not lead to changes in behavior or cell proliferation/survival in the brain. Together, these data indicate a potential role of cell proliferation/survival in selected brain areas on different behaviors between vole species with distinct life strategies.

Covering the Cover

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Role of zonula occludens in reduced epithelial cell proliferation by hypertonic stress

In addition to their role as epithelial barriers, tight junctions (TJs) function as dynamic signaling reservoirs involved in regulating the proliferative potential of epithelial cells. Hypertonic stress reduces cell proliferation and alters cell morphology. Here, we examined whether TJ zonula occludens (ZO) modulate cell proliferation under conditions of chronic hypertonic stress. As revealed by microscopy analysis and FACS, proliferation of spontaneously immortalized renal principal collecting duct (CD) mCCDcl1 cells is strongly dependent on cell confluency when grown under isotonic conditions (300 mOsmol/kg). Low levels of cell proliferation were accompanied by a reduced S phase and increased G0/G1 phase. Real‐Time PCR and Western blot analysis showed that ZO‐3 abundance increased with cell confluency while ZO‐1 and ZO‐2 abundance was unaffected. Confocal microscopy analysis revealed that expression of all three ZO isoforms at the cell surface increased with cell confluency. All observed changes were enhanced in slowly proliferating cells adapted to NaCl‐hypertonic medium (500 mOsmol/kg). Under isotonic conditions, knockdown of ZO‐1 and ZO‐2 by siRNA increased G1 phase, while knockdown of ZO‐3 promoted cell cycle progress. Similar results were obtained in cells habituated to hypertonic medium. These data reveal differential effects between ZO isoforms on renal CD cell proliferation. In addition, they indicate that ZO submembrane recruitment may influence slow cell proliferation under hypertonic conditions.

The growth and osteoclastogenic effects of fibroblasts isolated from keratocystic odontogenic tumor

To investigate the growth characteristics and effects on osteoclastogenesis in fibroblasts isolated from keratocystic odontogenic tumor (KCOT) fibrous capsule. Fibroblasts isolated from KCOT fibrous capsule and normal gingival mucosa were cultured in vitro. Their colony-forming units and proliferative activity were investigated, and the osteoclastogenic effects were also observed by a co-culture system with osteoclast precursor cell line Raw264.7. The mRNA of several genes related to bone resorption (IL-6, VEGF, COX-2, and M-CSF) was analyzed by real-time PCR. Keratocystic odontogenic tumor fibroblasts developed fewer CFU and had longer population doubling time than gingival fibroblasts (P < 0.05). In contrast to gingival fibroblasts, KCOT fibroblasts expressed less IL-6, COX-2, and M-CSF (P < 0.05); however, the Raw264.7 co-cultured with KCOT fibroblasts developed more osteoclast-like cells and expressed higher level of nfatc1 than that co-cultured with gingival fibroblasts. Increased COX-2 expression and VEGF expression were detected in KCOT fibroblasts and Raw264.7 co-culture system (P < 0.05). Although KCOT fibroblasts showed lower level of cell proliferation than gingival fibroblasts, higher osteoclastogenic ability was detected when co-cultured with Raw264.7. These results suggest that the cell-cell interaction in the co-culture system, possibly by increasing COX-2 and VEGF expression, may be responsible for the increased osteoclastogenic effects of KCOT fibroblasts.

The RhoA-ROCK-PTEN pathway as a molecular switch for anchorage dependent cell behavior

The proliferation of anchorage-dependent cells of mesenchymal origin requires the attachment of the cells to substrates. Thus, cells that are poorly attached to substrates exhibit retarded cell cycle progression or apoptotic death. A major disadvantage of most polymers used in tissue engineering is their hydrophobicity; hydrophobic surfaces do not allow cells to attach firmly and, therefore, do not allow normal proliferation rates. In this study, we investigated the molecular mechanism underlying the reduced proliferation rate of cells that are poorly attached to substrates. There was an inverse relationship between the activity of the small GTPase RhoA (RhoA) and the cell proliferation rate. RhoA activity correlated inversely with the strength of cell adhesion to the substrates. The high RhoA activity in the cells poorly attached to substrates caused an increase in the activity of Rho-associated kinase (ROCK), a well-known effector of RhoA that upregulated the activity of phosphatase and tensin homolog (PTEN). The resulting activated PTEN downregulated Akt activity, which is essential for cell proliferation. Thus, the cells that were poorly attached to substrates showed low levels of cell proliferation because the RhoA-ROCK-PTEN pathway was hyperactive. In addition, RhoA activity seemed to be related to focal adhesion kinase (FAK) activity. Weak FAK activity in these poorly attached cells failed to downregulate the high RhoA activity that restrained cell proliferation. Interestingly, reducing the expression of any component of the RhoA-ROCK-PTEN pathway rescued the proliferation rate without physico-chemical surface modifications. Based on these results, we suggest that the RhoA-ROCK-PTEN pathway acts as a molecular switch to control cell proliferation and determine anchorage dependence. In cells that are poorly attached to substrates, its inhibition is sufficient to restore cell proliferation without the need for physico-chemical modification of the material surface.

NNOS deficiency-induced cell proliferation depletes the neurogenic reserve

The consequences of nitric oxide synthase (NOS) gene knockout on proliferation, survival and differentiation of neuronal precursors in the subgranular (SGZ) and subventricular (SVZ) zones were analyzed. Comparative studies were performed in neonatal, adult and old (18-month) wild-type (WT), nNOS, eNOS, and iNOS knockout (KO) mice. Effects on brain cell proliferation were studied by sacrificing animals at 24h after injecting BrdU, while effects on survival and differentiation of dividing brain cells were studied by sacrificing other animals at three weeks after injections and double immunostaining with cell phenotype-specific antibodies. In the neonatal SGZ, cell proliferation was higher than at any other age, with a significantly decreased level in eNOS-KO mice. In the neonatal SVZ, cell proliferation in each of the three NOS-KO strains was significantly lower than in WT. In the adult, in both the SGZ and SVZ, all strains showed lower levels of cell proliferation than in neonates. Thereby, the significant highest cell proliferation was found in the SGZ and SVZ of nNOS-KO mice. In the SGZ and SVZ of old mice, in each strain, BrdU-positive cell counts were further reduced from adult levels, whereby cell proliferation of nNOS-KO mice attained the most massive reduction (in the SGZ almost to zero). In adult animals sacrificed 21 days after BrdU injections, values of BrdU-/NeuN-positive cells in all knockout animals were the same as WT, indicating that the initial cell proliferation changes were not sustained or translated into neuronal differentiation. The effect of nNOS-KO, inducing cell proliferation only temporarily, consists with the concept that neuronal stem cells have a finite proliferation capacity.

Ascl3 knockout and cell ablation models reveal complexity of salivary gland maintenance and regeneration

Expression of the transcription factor, Ascl3, marks a population of adult progenitor cells, which can give rise to both acinar and duct cell types in the murine salivary glands. Using a previously reported Ascl3EGFP-Cre/+ knock-in strain, we demonstrate that Ascl3-expressing cells represent a molecularly distinct, and proliferating population of progenitor cells located in salivary gland ducts. To investigate both the role of the Ascl3 transcription factor, and the role of the cells in which it is expressed, we generated knockout and cell-specific ablation models. Ascl3 knockout mice develop smaller salivary glands than wild type littermates, but secrete saliva normally. They display a lower level of cell proliferation, consistent with their smaller size. In the absence of Ascl3, the cells maintain their progenitor function and continue to generate both acinar and duct cells. To directly test the role of the progenitor cells, themselves, in salivary gland development and regeneration, we used Cre-activated expression of diphtheria toxin (DTA) in the Ascl3-expressing (Ascl3+) cell population, resulting in specific cell ablation of Ascl3+ cells. In the absence of the Ascl3+ progenitor cells, the mice developed morphologically normal, albeit smaller, salivary glands able to secrete saliva. Furthermore, in a ductal ligation model of salivary gland injury, the glands of these mice were able to regenerate acinar cells. Our results indicate that Ascl3+ cells are active proliferating progenitors, but they are not the only precursors for salivary gland development or regeneration. We conclude that maintenance of tissue homeostasis in the salivary gland must involve more than one progenitor cell population.

Exogenous neuropeptide Y promotes in vivo hippocampal neurogenesis

Adult neurogenesis mainly occurs in two brain regions, the subventricular zone and the dentate gyrus (DG) of the hippocampus. Neuropeptide Y (NPY) is widely expressed throughout the brain and is known to enhance in vitro hippocampal cell proliferation. Mice lacking either NPY or the Y1 receptor display lower levels of cell proliferation, thereby suggesting a role for NPY in basal in vivo neurogenesis. Here, we investigated whether exogenous NPY stimulates DG progenitors proliferation in vivo. We show that intracerebroventricular administration of NPY increases DG cell proliferation and promotes neuronal differentiation in C57BL/6 adult mice. In these mice, the proliferative effect of NPY is mediated by the Y1 and not the Y2 receptor, as a Y1 ([Leu(31) ,Pro(34) ]), but not a Y2 (NPY(3-36) ), receptor agonist enhanced proliferation. In addition, no NPY-induced DG cellular proliferation is observed following NPY injection when coadministered with a Y1 antagonist or in the Y1 receptor knockout mouse. These results are in line with data obtained in Y1(-/-) mice, demonstrating that NPY regulates in vivo hippocampal neurogenesis. © 2010 Wiley-Liss, Inc.

Two-fold elevation of expression of FoxM1 transcription factor in mouse embryonic fibroblasts enhances cell cycle checkpoint activity by stimulating p21 and Chk1 transcription

Forkhead Box M1 (FoxM1) transcription factor regulates expression of cell cycle effective genes and is stabilized by checkpoint kinase 2 (Chk2) to stimulate expression of DNA repair enzymes in response to DNA damage. This study intended to test whether FoxM1 is involved in cell cycle checkpoint pathways. Analysis of senescence and cell proliferation in FoxM1 transgenic (TG) mouse embryonic fibroblasts (MEFs) with 2-fold elevation of FoxM1, and overexpression or knockdown of FoxM1 in an inducible FoxM1 expression cell line, or FoxM1 siRNA. Chromatin immunoprecipitation (ChIP), electrophoretic mobility shift assays (EMSA), and cotransfection to determine FoxM1 transcription targets, as well as RNase protection assays and western blot analysis, were performed. Two-fold elevation of FoxM1 in FoxM1-TG-MEFs resulted in low levels of cell proliferation and increase in permanent cell cycle arrest at early passages (from passage 6 to 9). These phenotypes correlated with increased phosphorylation of p53 on Ser15, elevated expression of cell cycle inhibitor p21 and Chk1 at passage 3. FoxM1 was stabilized in response to DNA damage in MEFs and FoxM1 overexpression induced p21. Knockdown of FoxM1 resulted in decrease in Chk1. ChIP, EMSA and cotransfection assays confirmed that FoxM1 stimulated promoters of p21 and Chk1. Chk1 and p21 are direct transcription targets of FoxM1 and FoxM1 participates in transcriptional responses to stress in normal cells.

Estradiol does not influence strategy choice but place strategy choice is associated with increased cell proliferation in the hippocampus of female rats

Adult neurogenesis occurs in the hippocampus of most mammals. While the function of adult hippocampal neurogenesis is not known, there is a relationship between neurogenesis and hippocampus-dependent learning and memory. Ovarian hormones can influence learning and memory and strategy choice. In competitive memory tasks, higher levels of estradiol shift female rats towards the use of the place strategy. Previous studies using a cue-competition paradigm find that 36% of male rats will use a hippocampus-dependent place strategy and place strategy users had lower levels of cell proliferation in the hippocampus. Here, we used the same paradigm to test whether endogenous or exogenous ovarian hormones influence strategy choice in the cue-competition paradigm and whether cell proliferation was related to strategy choice. We tested ovariectomized estradiol-treated (10 μg of estradiol benzoate) or sham-operated female rats on alternating blocks of hippocampus-dependent and hippocampus-independent versions of the Morris water task. Rats were then given a probe session with the platform visible and in a novel location. Preferred strategy was classified as place strategy (hippocampus-dependent) if they swam to the old platform location or cue strategy (hippocampus-independent) if they swam to the visible platform. All groups showed a preference for the cue strategy. However, proestrous rats were more likely to be place strategy users than rats not in proestrus. Female place strategy users had increased cell proliferation in the dentate gyrus compared to cue strategy users. Our study suggests that 78% of female rats chose the cue strategy instead of the place strategy. In summary the present results suggest that estradiol does not shift strategy use in this paradigm and that cell proliferation is related to strategy use with greater cell proliferation seen in place strategy users in female rats.

Hippocampus-dependent strategy choice predicts low levels of cell proliferation in the dentate gyrus

Neurogenesis continues to occur throughout life in the mammalian hippocampus. Previous research has suggested that the production of new neurons in the hippocampus during adulthood may be related to hippocampus-dependent learning and memory. However, the exact relationship between adult neurogenesis and learning and memory remains unclear. Here we investigated whether learning strategy selection is related to cell proliferation or to survival of new neurons in the hippocampus of adult male rats. We trained rats on alternating blocks of hippocampus-dependent (hidden platform) and hippocampus-independent (visible platform) versions of the Morris water task with the platform always in the same position. Following training, rats were given a probe session during which the platform was visible and in a novel location. Preferred strategy was determined by observing the initial swim path. Rats were classified as place strategy (hippocampus-dependent) users if they swam to the old platform location. Cue strategy (hippocampus-independent) users were classified as those rats that swam initially to the visible platform. Our results indicate that rats that preferentially used a place strategy had significantly lower cell proliferation than cue strategy users. However, there was no significant difference in cell survival or number of immature neurons between strategy user groups. These results suggest that low levels of cell proliferation in the dentate gyrus may be conducive or coincident with more efficient memory processing in the hippocampus.

Human bone marrow-derived mesenchymal stem cells and osteoblast differentiation on titanium with surface-grafted chitosan and immobilized bone morphogenetic protein-2

Circulating progenitor cells are known to home to various organs to repair injured tissues or to routinely replace old cells and maintain tissue integrity. Similarly, circulating progenitor bone cells can possibly home to a bone implant, differentiate, and eventually osteointegrate with the prosthesis. Osteointegration of bone cells with the prosthesis can help to reduce the risk of implant failure due to constant movement between bone tissue and implant surface. In this study, we aim to investigate if immobilized bone morphogenetic protein-2 (BMP2) on chitosan-grafted titanium substrate (Ti-CS-BMP2) will enhance bone marrow-derived mesenchymal stem cell (BMMSC) adhesion onto the substrate surface and further induce their differentiation into osteoblasts. The results show that our Ti-CS-BMP2 substrate is able to retain adsorbed BMP2, and is capable of slow release of this growth factor. Despite the lesser number of BMMSCs initially attached onto the Ti-CS-BMP2 substrates and consequently the lower level of cell proliferation, Ti-CS-BMP2 cells had the highest level of ALP activity. RT-PCR results show that Ti-CS-BMP2 cells had a relatively higher level of transcription activity of Runx2, compared with that of bone cell-derived osteoblasts (BC-OB), an indication that the BMMSCs were actively differentiating into osteoblasts. Finally, alizarin red staining reveals the presence of calcium deposits in the differentiated cells. Hence our Ti-CS-BMP2 substrates possess an osteoconductive effect and can possibly be used to fabricate bone implants that can osteointegrate with host bone tissue.

Involvement of PKC‐ι in glioma proliferation

Atypical protein kinase C-iota (PKC-iota) protects cells against apoptosis and may play a role in cell proliferation. However, in vivo, the status and function of PKC-iota in human normal brain tissue, gliomas, benign and malignant meningiomas as well as its in vitro status in proliferating and confluent glioma cells, remains unknown. The objectives of our research were to determine whether expression of PKC-iota is altered either in gliomas or in benign and malignant meningiomas, compared to normal brain. In addition, we wished to establish the expression of PKC-iota in proliferating plus in cell cycle-arrested glioma cell lines, as well as the relationship between PKC-iota siRNA on PKC-iota protein content and cell proliferation. Western blot analyses for PKC-iota were performed on 12 normal brain biopsies, 15 benign meningiomas, three malignant meningiomas and three gliomas. Results demonstrated no (n = 9) or very weak (n = 3) detection of PKC-iota in normal brain tissue. In comparison, PKC-iota was robustly present in the majority of the benign meningiomas. Similarly, PKC-iota was abundant in all malignant meningiomas and gliomas. Western blotting for PKC-iota in confluent or proliferating glioma cell lines depicted substantial quantities of PKC-iota in proliferating T98G and U-138MG glioma cells. In contrast, confluent cells had either 71% (T98G) or 21% (U-138MG) less PKC-iota than proliferating cells. T98 and U-138 MG glioma cells treated with 100 nm PKC-iota siRNA had lower levels of cell proliferation compared to control siRNA-A and complete down-regulation of PKC-iota protein content. These results support the concept that presence of PKC-iota may be required for cell proliferation to take place.