Evidence that brain-derived neurotrophic factor is required for basal neurogenesis and mediates,in part,the enhancement of neurogenesis by dietary restriction in the hippocampus of adult mice

To determine the role of brain-derived neurotrophic factor (BDNF) in the enhancement of hippocampal neurogenesis resulting from dietary restriction (DR), heterozygous BDNF knockout (BDNF +/-) mice and wild-type mice were maintained for 3 months on DR or ad libitum (AL) diets. Mice were then injected with bromodeoxyuridine (BrdU) and killed either 1 day or 4 weeks later. Levels of BDNF protein in neurons throughout the hippocampus were decreased in BDNF +/- mice, but were increased by DR in wild-type mice and to a lesser amount in BDNF +/- mice. One day after BrdU injection the number of BrdU-labeled cells in the dentate gyrus of the hippocampus was significantly decreased in BDNF +/- mice maintained on the AL diet, suggesting that BDNF signaling is important for proliferation of neural stem cells. DR had no effect on the proliferation of neural stem cells in wild-type or BDNF +/- mice. Four weeks after BrdU injection, numbers of surviving labeled cells were decreased in BDNF +/- mice maintained on either AL or DR diets. DR significantly improved survival of newly generated cells in wild-type mice, and also improved their survival in BDNF +/- mice, albeit to a lesser extent. The majority of BrdU-labeled cells in the dentate gyrus exhibited a neuronal phenotype at the 4-week time point. The reduced neurogenesis in BDNF +/- mice was associated with a significant reduction in the volume of the dentate gyrus. These findings suggest that BDNF plays an important role in the regulation of the basal level of neurogenesis in dentate gyrus of adult mice, and that by promoting the survival of newly generated neurons BDNF contributes to the enhancement of neurogenesis induced by DR.     

Alterations in gastric mucosal lineages induced by acute oxyntic atrophy in wild-type and gastrin-deficient mice

In addition to their role in gastric acid secretion, parietal cells secrete a number of growth factors that may influence the differentiation of other gastric lineages. Indeed, oxyntic atrophy is considered the most significant correlate with increased risk for gastric adenocarcinoma. We studied the alterations in gastric mucosal lineages elicited by acute oxyntic atrophy induced by treatment of C57BL/6 and gastrin-deficient mice with the parietal cell protonophore [S-(R*,S*)]-N-[1-(1,3-benzodioxol-5-yl)butyl]-3,3-diethyl-2-[4-[(4-methyl-1-piperazinyl)carbonyl]phenoxy]-4-oxo-1-azetidinecarboxamide (DMP-777). In both wild-type and gastrin knockout mice, DMP-777 elicited the rapid loss of parietal cells within 2 days of treatment. In wild-type mice, oxyntic atrophy was accompanied by a rapid increase in 5-bromo-2'-deoxyuridine-labeled proliferative cells and attendant increase in surface cell numbers. However, gastrin knockout mice did not demonstrate significant foveolar hyperplasia and showed a blunted proliferative response. After 7 days of treatment in wild-type mice, a second proliferative population emerged at the base of fundic glands along with the development of a mucous cell metaplasia expressing TFF2/spasmolytic polypeptide (SPEM). However, in gastrin knockout mice, SPEM expressing both TFF2 mRNA and protein developed after only 1 day of DMP-777 treatment. In wild-type mice, all changes induced by DMP-777 were reversed 14 days after cessation of treatment. In gastrin-deficient mice, significant SPEM was still present 14 days after the cessation of treatment. The results indicate that foveolar hyperplasia requires the influence of gastrin, whereas SPEM develops in response to oxyntic atrophy independent of gastrin, likely through transdifferentiation of chief cells.     

Progenitor cell cycling during hair cell regeneration in the vestibular and auditory epithelia of the chick

We investigated nucleotide-labeling patterns during ongoing hair cell regeneration in the avian vestibular epithelium and during drug-induced regeneration in the avian auditory epithelium. For utricle experiments, post-hatch chicks received an injection of bromodeoxyuridine (BrdU) and were allowed to survive from 2 hours to 110 days after the injection. Utricles were fixed and immunoreacted to detect BrdU. The number of BrdU-labeled nuclei in the hair cell and support cell layers of the utricular sensory epithelium changes significantly between 2 hours and 110 days post-BrdU. At 2 hours, most labeled cells are isolated, while by 5–10 days, the majority of labeled cells are organized in pairs that are most frequently composed of a hair cell and a support cell. Pairs of labeled cells are seen as late as 110 days. Clusters of more than 3 labeled cells are uncommon at all time-points. The total number of labeled cells increases approximately 1.5-fold between 5 and 60 days post-BrdU. This increase is due primarily to a rise in the number of labeled support cells, and it is likely that it represents additional rounds of division by a subset of cells that were labeled at the time of the BrdU injection. There is a significant decrease in labeled nuclei in the hair cell layer between 60 and 110 days post-BrdU, suggesting that hair cells die during this period. To investigate support cell recycling in the drug-damaged auditory epithelium, we examined nucleotide double labeling after separate injections of BrdU and tritiated thymidine. A small number of support cells that incorporate BrdU administered at 3 days post-gentamicin treatment also label with tritiated thymidine administered between 17 and 38 hours later. We conclude that a small population of support cells recycles during regeneration in both the normal utricle and the drug-damaged basilar papilla.     

Alternative ultrastructural localization of Dolichos biflorus lectin binding sites in proton secreting parietal cells of mice

High amount of N-acetyl-D-galactosamine specific lectin binding sites were detected on the canalicular membranes of human parietal cells. Our present model investigations on mice showed that the intracellular distribution of the terminal N-acetyl-D-galactosamine containing glycoprotein highly depends on the actual functional state of the parietal cells. In the normal gastric mucosa 40%-60% of parietal cells react positively after staining with horseradish peroxidase or biotin labelled Dolichos biflorus lectin. Ultrastructurally lectin binding sites occur mainly on the basolateral membrane infoldings in fed animals, while they are present exclusively on the canalicular membranes of fasting mice, suggesting that the alternative appearance of lectin binding sites on the opposite membrane areas of parietal cells is tightly coupled to their main function, to H+ secretion.     

IGF-1对不同年龄缺血性脑损伤大鼠神经发生的影响

目的:检测胰岛素样生长因子-1(IGF-1)对青年和老年大鼠局灶脑缺血后神经发生及其后细胞生存的影响.方法:健康雄性SD青年鼠(3-4个月)和老年鼠(1年)随机分组,侧脑室注入IGF-1,1天后进行大鼠大脑中动脉阻塞(MCAO),对照组由生理盐水取代.采用BrdU标记方法鉴定MCAO后7d和28d的增殖细胞.BrdU于MCAO后第6d由腹腔注入.免疫组化法检测7天后BrdU、PSA-NCAM标记细胞和28天后BrdU、BrdU/MAP2双标细胞.结果:老年组中BrdU阳性细胞的数目7d后较对照组增加5.1倍;青年纽中BrdU阳性细胞的数目7d后较对照组增加5.5倍.28d后,BrdU阳性细胞的残留率在青年IGF-1处理组和老年IGF-1处理组中分别是79.2%和75.1%,分别相对于对照组的77.1%和52.3%.老年组中PSA-NCAM阳性细胞的数目7d后较对照组增加3.2倍;青年组中PSA-NCAM阳性细胞的数目7d后较对照组增加3.7倍.28d后,BrdU/MAP2阳性细胞在青年IGF-1处理组较对照组增加7.0倍,在老年IGF-1处理组较对照组增加4.9倍.结论:此结果提示局部应用IGF-1进行缺血前预处理,在青年鼠和老年鼠中均能诱导神经发生,且在老年鼠中能明显提高神经发生后的增殖细胞的生存率和向神经元分化的能力.这一研究结果将有助于研究IGF-1在中老年脑损伤病人中的治疗性应用.     

自体骨髓单核细胞移植在肝纤维化兔肝脏内的定植能力

目的建立四氯化碳诱导的兔肝纤维化动物模型,观察体外分离标记的自体骨髓单核细胞（ABM-MNCs）经肠系膜上静脉自体移植至肝纤维化区及周边区后的存活、定植状况。方法将40只普通级日本大耳家兔随机分为细胞移植组和对照组各20只,实验组腹腔注射40%CCl4橄榄油溶液建立肝纤维化模型,对照组腹腔注射等量生理盐水。细胞移植组于模型稳定后自体髂骨处抽取骨髓,采用氯化氨红细胞溶解法分离得到单核细胞,以5溴-2脱氧尿嘧啶核苷（BrdU）标记体外ABM-MNCs及鉴定;分离培养ABM-MNCs,将3×10^9个ABM-MNCs经肠系膜上静脉回输体内,对照组回输等量生理盐水,移植前、移植后3、7、14、21 d分别取肝组织固定,进行免疫组织化学检测。结果BrdU体外标记ABM-MNCs的免疫组织化学表现示：20μmol/L BrdU孵育ABM-MNCs 72 h的阳性标记率达95%;肝组织20μmol/L BrdU免疫组化染色切片显示：自体骨髓单核细胞移植后第3天,肝小叶中央静脉周围BrdU染色阳性,随着时间的推移,阳性染色逐渐增强,并逐步向肝组织内部延伸。阳性染色主要分布于肝组织汇管区周围组织,而对照组BrdU染色则阴性。结论ABM-MNCs经肠系膜上静脉移植后,可在纤维化区及周边区存活,定植。     

Role of hyaluronan and CD44 in reactive oxygen species-induced mucus hypersecretion

5-bromo-2-deoxyurudine (BrdU) can be used as a methodological tool for in vivo investigations following in vitro prelabeling of isolated stem cells for subsequent cell tracking within the recipient host. The objective of this study was to determine how useful BrdU may be as a labeling modality for adipose derived stem cells (ASC) by examining BrdU toxicity, BrdU intracellular stability, and potential effects on ASC differentiation. Porcine and human ASC (pASC and hASC, respectively) were labeled with BrdU at 5 or 10 μM for 2, 6, 24, and 48 h. BrdU toxicity and stability over time in monolayer cultures, in 3-D collagen scaffolds implanted to a porcine model and after thawing from long-term storage were evaluated by MTT assays and immunohistochemistry. ASC differentiation was evaluated by Oil Red O staining. BrdU was not cytotoxic at all tested concentrations and incubation times. BrdU color intensity within each cell and the number of ASC labeled with BrdU decreased as a function of both incubation time and BrdU concentrations. Labeling intensities decreased over time and were undetectable after 6 passages for pASC and 4 passages for hASC. In 3-D scaffolds, BrdU-labeled ASC were identifiable after 90 days of in vitro cultures and for 30 days in a porcine model. BrdU did not prevent preadipocyte differentiation and BrdU labeling was still detectable after subsequent thawing after long-term storage of ASC. BrdU is an excellent candidate reagent to label and track ASC that will allow distinction between BrdU-labeled donor cells and host cells. The data provides a foundation for conducting future tissue engineering projects using BrdU-labeled ASC.     

Identification of putative bovine mammary epithelial stem cells by their retention of labeled DNA strands

Stem cells appear to retain labeled DNA for extended periods because of their selective segregation of template DNA strands during mitosis. In this study, proliferating cells in the prepubertal bovine mammary gland were labeled using five daily injections of 5-bromo-2-deoxyuridine (BrdU). Five weeks later, BrdU-labeled mammary epithelial cells were still evident. The percentage of BrdU-labeled epithelial cells was greatest in the lower region of the mammary gland, near the gland cistern, and was decreased toward the periphery of the parenchymal region, where the ducts were invading the mammary fat pad. Increased numbers of BrdU-labeled epithelial cells in basal regions of the gland are likely a consequence of decreased proliferation rates and increased cell cycle arrest in this area. In peripheral regions of mammary parenchyma, the percentage of heavily labeled epithelial cells averaged 0.24%, a number that is consistent with estimates of the frequency of stem cells in the mouse mammary gland. Epithelial label-retaining cells seemingly represent a slowly proliferating population of cells, as 5.4% of heavily labeled cells were positive for the nuclear proliferation antigen Ki67. Because epithelial label-retaining cells contain estrogen receptor (ER)-negative and ER-positive cells, they apparently comprise a mixed population, which I suggest is composed of ER-negative stem cells and ER-positive progenitors. Continuing studies will address the usefulness of this technique to identify bovine mammary stem cells and to facilitate studies of stem cell biology.     

Detection of in situ mitotic activity of dendritic epidermal T-cells by BrdU labeling.

We analyzed mitotic dendritic epidermal T-cells (DETC) in the epidermis of C3H/He (Thy-1.2+) mice, using double immunoenzymatic labeling. Ear skin was incubated with 100 microM bromodeoxyuridine (BrdU) for 5 hr and then either directly studied or cultured for an additional 12 hr in BrdU-free medium. After BrdU labeling, with or without additional culture, epidermal sheets were obtained by ethylenediaminetetraacetic acid separation. The epidermal specimens were immunostained by the peroxidase method to visualize nuclear BrdU and then by the biotin-streptavidin-alkaline phosphatase method for surface Thy-1.2 antigen. In specimens processed immediately after BrdU labeling, a mean 3.0% of all basal cells were labeled with BrdU and a mean 1.1% of the BrdU-labeled cells were also positive for Thy-1.2. Moreover, a mean 2.1% of the DETC had incorporated BrdU. BrdU-labeled DETC had a variety of appearances; they were dendritic and round in the BrdU-treated specimens, while oval and paired cells were also found in the specimens after additional culture. These morphological changes of BrdU-labeled DETC demonstrate that resident DETC can become mother cells undergoing mitosis through the retraction of their dendrites, and it appears that DETC divide at a relatively high rate, i.e., up to 10% of the DETC may enter the S-phase of the cell cycle every 24 hr.     

In vitro bromodeoxyuridine labeling of nuclei: application to myotube hybridization

Rat myoblast nuclei were labeled with various concentrations of bromodeoxyuridine (BrdU), an analogue of thymidine, for 24 or 48 hr. Almost every myoblast was labeled with BrdU at concentrations between 10(-7) M and 10(-5) M. When the cells were labeled with 0.5 microM or more, the percentage of labeled cells remained over 90% and 80% at 2 and 5 days, respectively. However, when the cells were labeled with BrdU concentration lower than 10(-7) M the percentage of labeled nuclei decreased more rapidly with time. The BrdU-labeled cells were mixed with an unlabeled population to determine whether their capacity to fuse was reduced. At a BrdU concentration of 0.5 x 10(-6) M, labeled myoblasts fused to a similar extent as unlabeled myoblasts, and a high percentage of marked cells were still perceptively labeled after 5 days. In contrast, the fusion capacity of myoblasts incubated with more than 10(-6) M BrdU was inhibited after only few rounds of DNA synthesis. These myoblasts were eventually able to fuse, however, when the BrdU diminished in the DNA due to cell division. These results indicate that labeling with BrdU at a concentration of 0.5 x 10(-6) M and an incorporation time of 48 hr is optimal to obtain perceptible immunocytochemical staining without affecting myoblast fusion. Such BrdU immunolabeling could be used as a nuclear marker for hybridization studies.     

Solitary chemoreceptor cell proliferation in adult nasal epithelium

Nasal trigeminal chemosensitivity in mice and rats is mediated in part by solitary chemoreceptor cells (SCCs) in the nasal epithelium (Finger et al., 2003). Many nasal SCCs express the G-protein α-gustducin as well as other elements of the bitter-taste signaling cascade including phospholipase Cβ2, TRPM5 and T2R bitter-taste receptors. While some populations of sensory cells are replaced throughout life (taste and olfaction), others are not (hair cells and carotid body chemoreceptors). These experiments were designed to test whether new SCCs are generated within the epithelium of adult mice. Wild type C57/B6 mice were injected with the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) to label dividing cells. At various times after injection (1–40 days), the mice were perfused with 4% paraformaldehyde and prepared for dual-label immunocytochemistry. Double labeled cells were detected as early as 3 days post BrdU injection and remained for as long as 12 days post-injection suggesting that SCCs do undergo turnover like the surrounding nasal epithelium. No BrdU labeled cells were detected after 24 days suggesting relatively rapid replacement of the SCCs.     

Supporting cell proliferation after hair cell injury in mature guinea pig cochlea in vivo

In cold-blooded animals, lost sensory hair cells can be replaced via a process of regenerative cell proliferation of epithelial supporting cells. In contrast, in mammalian cochlea, receptor (hair) cells are believed to be produced only during embryogenesis; after maturity, sensory or supporting cell proliferation or regeneration are thought to occur neither under normal conditions nor after trauma. Using bromodeoxyuridine (BrdU) as a proliferation marker, we have assessed cell proliferation activity in the mature organ of Corti in the cochlea of young guinea pigs following severe damage to the outer hair cells induced by kanamycin sulfate and ethacrynic acid. Although limited, we have found BrdU-labeled nuclei in the regions of Deiters cells when BrdU is given for 3 days or longer. When BrdU is given for 10 days, at least one labeled nucleus can be observed in the organ of Corti in approximately half of the ears; proliferating cells typically appear as paired daughters, with one nucleus being displaced away from the basement membrane to the position expected of the hair cells. Double-staining with antibodies to cytokeratin, vimentin, and p27 have shown that the BrdU-labeled nuclei are located in cells phenotypically similar to Deiters cells. Most of the uptake of BrdU occurs 3–5 days following ototoxic insult, and the number of BrdU-labeled cells does not decrease until 30 days following insult. These findings indicate that Deiters cells in the mature mammalian cochlea maintain a limited competence to re-enter the cell cycle and proliferate after hair cell injury, and that they can survive at least for 1 month.This work was supported by the Ministry of Health, Labour, and Welfare, Japan (grants 12120101, 15110201) and by the Ministry of Education, Culture, Sports, Science, and Technology, Japan (grant 13470357) to T.Y.     

Selective binding of Dolichos biflorus agglutinin to L3T4-, Lyt-2- thymocytes. Expression of terminal alpha-linked N-acetyl-D-galactosamine residues defines a subpopulation of fetal and adult murine thymocytes

Using histochemical and immunocytochemical techniques, a lectin with nominal specificity for alpha-linked N-acetyl-D-galactosamine, Dolichos biflorus agglutinin (DBA), was found to preferentially label thymocytes with an L3T4-, Lyt-2- phenotype from fetal/newborn and adult mice. Through days 14 to 16 of gestation, virtually all thymocytes bound DBA, followed by a dramatic reduction of DBA labeling during the last 4 days of gestation, reaching adult levels of about 2 to 4% of total thymocytes. At later stages of fetal development, the DBA+ cells were confined to the subcapsular area of the thymus. This apparent loss of DBA+ cells was caused by an expansion of the thymocyte population not labeled with this lectin. Affinity purification of thymocyte cell surface components with insolubilized DBA indicated that virtually all of the lectin binding to fetal thymocytes was mediated by a 120-kDa glycoprotein. In addition to thymocytes, DBA also labeled about 5% of bone marrow cells from both normal or nude mice and a small population of spleen cells as well. These results suggest that this lectin may be useful to positively select for LT34-, Lyt-2- thymocytes, and, possibly, other immature populations within the T cell lineage.     

Purkinje cell fate in staggerer mutants: agenesis versus cell death

Staggerer (sg/sg) is an autosomal recessive mutation in an orphan nuclear hormone receptor gene, RORalpha, that causes a cell-autonomous, lineage-specific block in the development of the Purkinje cell. Purkinje cell number is reduced by about 75-90% in adult mutants, and many of the surviving cells are small and ectopically positioned. To determine whether Purkinje cell numbers are reduced owing to either agenesis or cell death, cohorts of Purkinje cells were labeled with the birth-date marker bromodeoxyuridine (BrdU) at embryonic day (E) 10.5 or E11.5. The total number of BrdU-labeled profiles was then compared between cerebella from wild-type mice, heterozygous staggerer, and staggerer mutants at E17.5 and postnatal day (P)5. There was no significant difference between sg/sg mutants and +/sg or +/+ controls in the number of BrdU-labeled profiles or in cerebellar volumes in the E17 embryos. By P5, however, cerebellar volume was significantly reduced in the sg/sg mutants compared to controls (p <.005) and the number of BrdU-labeled profiles was reduced by 33% following E11.5 BrdU injections (p <.02). The results suggest that Purkinje cell genesis is not affected by the staggerer mutation and that Purkinje cell loss begins some time after E17. RORalpha is highly expressed in Purkinje cells by E14, so the delay between initial RORalpha expression and sg/sg Purkinje cell loss suggests that the staggerer mutation does not directly cause Purkinje cell death.     

The timing of alpha-gustducin expression during cell renewal in rat vallate taste buds

The G protein subunit alpha-gustducin is expressed in a subset of light (Type II) but not in dark (Type I) cells in rat vallate taste buds. The thymidine analogue 5-bromo-2'-deoxyuridine (BrdU) is incorporated into DNA during the S-phase of the cell cycle and can be used to determine the time of origin of a cell. In this study, 31 rats were injected with BrdU (50 mg/kg i.p.) and perfused at various times, from 2.5 to 10.5 days, following BrdU administration. Vallate papillae were embedded in polyester wax, cut into 4 microm transverse sections, and characterized with antibodies to BrdU and alpha-gustducin. Sections were processed for indirect immunofluorescence or with an immunoperoxidase procedure. From immunoperoxidase material on 21 rats, counts of alpha-gustducin- and BrdU-labeled cells were obtained from 300-800 taste bud profiles at each survival time; a total of 4122 taste bud profiles were examined. Cells with nuclei immunoreactive for BrdU occurred within the taste buds at 2.5 days and double-labeled cells were clearly evident at 3.5 days; a small number of double-labeled cells were seen as early as 2.5 days. Double-labeled cells reached a peak at 6.5 days and did not decline significantly by 10.5 days. Cells labeled for BrdU but not alpha-gustducin peaked at 5.5 days and showed a significant decline by 8.5 days. These latter cells included light cells not expressing alpha- gustducin and dark cells, which have previously been shown to have a shorter life span than light cells. These data suggest that expression of alpha-gustducin appears very early in a cell's life span and that these cells are longer lived than many of the cells that do not express this G protein.      

Neurogenesis in the vomeronasal epithelium of adult rats: evidence for different mechanisms for growth and neuronal turnover

The pattern of cell migration during neuronal turnover in the vomeronasal sensory epithelium (VN-SE) is controversial. In mice, proliferating cells were detected at the edges and were described as migrating to the center of the VN-SE. In rats, in addition to proliferating cells at the margins of the epithelium, dividing cells are also present along the entire basal lamina of the VN-SE. In marsupials, dividing cells have also been observed in the margins and in the center of the VN-SE, the latter of which migrate vertically and become neurons. To investigate whether the process of neuronal turnover in placental mammals consists of horizontal and/or vertical migration, and whether or not this process is common to mammals, adult rats were injected with bromodeoxyuridine (BrdU) and allowed to survive for different periods of time. The distribution of BrdU-labeled cells in the horizontal and vertical dimension of the VN-SE was analyzed as a function of time. Both horizontal and vertical migrations of BrdU-labeled cells were detected. Since cells in the center of the VN-SE migrate vertically, and, as demonstrated by coexpression of markers of neuronal maturity and BrdU, become mature one day after undergoing mitosis, it is very likely that these cells participate in neuronal turnover. Conversely, because cells in the margins of the VN-SE stop migrating horizontally on day 14 before they have reached the center of the VN-SE, and since the VN-SE continues to grow during adulthood, it is likely that most of these latter cells constitute pools for growth.     

Chronic Helicobacter pylori infection results in gastric hypoacidity and hypergastrinemia in wild-type mice but vagally induced hypersecretion in gastrin-deficient mice

Helicobacter pylori infection is a causal factor of gastric cancer (which is associated with low gastric acid secretion) or duodenal ulcer (high acid secretion). Parietal cells and ECL cells in the stomach are controlled by gastrin, which plays a crucial role in the regulation of acid secretion. The present study was undertaken to identify a possible role of gastrin in determining the different responses of the parietal cells and ECL cells to chronic H. pylori infection. Wild-type (C57BL/6J) gastrin(+/+) mice and gastrin(-/-) knockout mice, generated through targeted gene disruption and backcrossed eight times to C57BL/6J, were infected with H. pylori for 9 months. The acid output was measured 4 h after pylorus ligation (known to cause vagal excitation). The gastric mucosa was examined by immunocytochemistry with antisera to alpha-subunit of H+/K(+)-ATPase for the parietal cells, and to histamine and vesicle monoamine transporter-2 for the ECL cells, and by quantitative electron microscopy. In infected gastrin(+/+) mice, the acid output and the percentage of secreting parietal cells (freely fed state) were 20-30% of the values in uninfected controls, while the density and ultrastructure of parietal cells were normal. The infected mice had hypergastrinemia and displayed hypertrophy and hyperplasia of ECL cells. Although uninfected gastrin(-/-) mice had lower the acid output than uninfected gastrin(+/+) mice, there was a higher acid output (approximately 3 times) in infected gastrin(-/-) mice than their uninfected homologues. The numbers of parietal cells and ECL cells remained unchanged in infected gastrin(-/-) mice. In conclusion, chronic H. pylori infection results to impaired parietal-cell function (acid hyposecretion), hypergastrinemia and hyperplasia of ECL cells in wild-type mice but leads to vagally induced hypersecretion in gastrin-deficient mice.     

IGF-1对缺血性脑损伤大鼠脑内神经发生的影响

目的:建立大鼠单侧局灶脑缺血模型,观察胰岛素样生长因子-1(IGF-1)对局灶脑缺血后的鼠脑神经发生及增殖后细胞生存的影响.方法:用健康雄性SD大鼠建立大脑中动脉阻塞(MCAO)模型,随机分成假手术组,缺血对照组和IGF-1治疗组.各组再按不同的治疗时间分为7d、14d、28d、42d组.免疫组化法观察BrdU、PSA-NCAM的变化,免疫双标法观察BrdU/PSA-NCAM、BrdU/MAP2和BrdU/GFAP的共同表达变化.结果:BrdU标记细胞和PSA-NCAM标记细胞计数均在缺血后第7d最多,分别是缺血对照组的4.0倍和1.8倍,是假手术组的9.9倍和5.4倍.BrdU和PSA-NCAM双标细胞在缺血发生后双侧SVZ和DG区可以检测到,于第7d计数最多,之后逐渐降低;而BrdU和MAP2以及BrdU和GFAP双标细胞却从第14d开始逐渐增多,直到第42d.随着BrdU/PSA-NCAM双标阳性表达的逐渐降低,BrdU/MAP2双标阳性表达逐渐增高,呈现此消彼涨的变化.结论:IGF-1侧脑室注射后,在早期(7d内)诱导了缺血性脑损伤后神经细胞的增殖;在中期(7d-14d)诱导了新生细胞的迁移;在后期(14d后)伴随着迁移的进行新生细胞逐渐发生了分化.     

Responses of BrdU label-retaining dental pulp cells to allogenic tooth transplantation into mouse maxilla

Recently, we demonstrated that a pulse of BrdU given to prenatal animals reveals the existence of slow-cycling long-term label-retaining cells (LRCs), putative adult stem or progenitor cells, which reside in the dental pulp. This study aims to clarify responses of LRCs to allogenic tooth transplantation into mouse maxilla using prenatal BrdU-labeling, in situ hybridization for osteopontin and periostin, and immunohistochemistry for BrdU, nestin, and osteopontin. The upper-right first molars were allografted in the original socket between BrdU-labeled and non-labeled mice or between GFP transgenic and wild-type mice. Tooth transplantation caused degeneration of the odontoblast layer, resulting in the disappearance of nestin-positive reactions in the dental pulp. On postoperative days 5–7, tertiary dentin formation commenced next to the preexisting dentin where nestin-positive odontoblast-like cells were arranged in the successful cases. In BrdU-labeled transplanted teeth, dense LRCs were maintained in the center of the dental pulp beneath the odontoblast-like cells including LRCs, whereas LRCs disappeared in the area surrounding the bone-like tissue. In contrast, LRCs were not recognized in the pulp chamber of non-labeled transplants through the experimental period. Tooth transplantation using GFP mice demonstrated that the donor cells constituted the dental pulp of the transplant except for endothelial cells and some migrated cells, and the periodontal tissue was replaced by host-derived cells except for epithelial cell rests of Malassez. These results suggest that the maintenance of BrdU label-retaining dental pulp cells play a role in the regeneration of odontoblast-like cells in the process of pulpal healing following tooth transplantation.     

Early determination and long-term persistence of adult-generated new neurons in the hippocampus of mice

New neurons are continually generated in the adult hippocampus, but the important question, whether adult neurogenesis is transient or leads to the lasting presence of new neurons, has not yet been answered. Dividing cells were labeled with bromodeoxyuridine (BrdU) and were investigated by means of immunofluorescence and confocal microscopy at several time-points 1 day to 11 months thereafter. BrdU-labeled neurons remained stable in number and in their relative position in the granule cell layer over at least 11 months. This finding implies that the addition of new neurons is not transient and that their final number and localization are determined early. By contrast, expression of immature markers beta-III-tubulin and doublecortin in BrdU-labeled cells, peaked early after division and was not detectable after 4 weeks. In transgenic mice expressing enhanced green fluorescent protein under the nestin promoter none of the BrdU/nestin-positive cells early after division expressed the mature marker NeuN, confirming that no dividing neurons were detected. These new data suggest that new neurons are recruited early from the pool of proliferating progenitor cells and lead to a lasting effect of adult neurogenesis.