c-kit marks late retinal progenitor cells and regulates their differentiation in developing mouse retina

Retinal progenitor cells are believed to display altered proliferation and differentiation during retinal development, suggesting that retinal progenitor cell populations are not homogeneous. However, the composition of progenitor cell populations is not known, due in part to the lack of known surface markers identifying distinct stages of retinal progenitor cells. We found a dramatic change in the expression profile of the cell surface antigens c-kit and stage-specific embryonic antigen-1 (SSEA-1) in retinal progenitor cells during development. While SSEA-1 was expressed early in development, c-kit expression peaked in late stage progenitor cells. The identification of these developmental markers enabled us to characterize distinct sub-populations of retinal progenitor cells. Progenitor cell subpopulations expressing either SSEA-1, c-kit, or both showed different proliferation and differentiation abilities. Although SSEA-1-positive cells were augmented by beta-catenin signaling, c-kit-positive cells were positively regulated by Notch signaling. Taken together, our data suggest that c-kit and SSEA-1 can be used to spatiotemporally differentiate retinal progenitor populations that have intrinsically distinct characteristics. Prolonged expression of c-kit by a retrovirus resulted in the promotion of proliferation and the appearance of nestin-positive cells in the presence of the c-kit ligand, stem cell factor (SCF). This suggests a role for c-kit, Notch, and the beta-catenin signaling network in retinal development.     

Prolonged elevated levels of c-kit+ progenitor cells after a myocardial infarction by beta 2 adrenergic receptor priming

Endogenous progenitor cells may participate in cardiac repair after a myocardial infarction (MI). The beta 2 adrenergic receptor (ß2-AR) pathway induces proliferation of c-kit+ cardiac progenitor cells (CPC) in vitro. We investigated if ß2-AR pharmacological stimulation could ameliorate endogenous CPC-mediated regeneration after a MI. C-kit+ CPC ß1-AR and ß2-AR expression was evaluated in vivo and in vitro. A significant increase in the percentage of CPCs expressing ß1-AR and ß2-AR was measured 7 days post-MI. Accordingly, 24 hrs of low serum and hypoxia in vitro significantly increased CPC ß2-AR expression. Cell viability and differentiation assays validated a functional role of CPC ß2-AR. The effect of pharmacological activation of ß2-AR was studied in C57 mice using fenoterol administered in the drinking water 1 week before MI or sham surgery or at the time of the surgery. MI induced a significant increase in the percentage of c-kit+ progenitor cells at 7 days, whereas pretreatment with fenoterol prolonged this response resulting in a significant elevated number of CPC up to 21 days post-MI. This increased number of CPC correlated with a decrease in infarct size. The immunofluorescence analysis of the heart tissue for proliferation, apoptosis, macrophage infiltration, cardiomyocytes surface area, and vessel density showed significant changes on the basis of surgery but no benefit due to fenoterol treatment. Cardiac function was not ameliorated by fenoterol administration when evaluated by echocardiography. Our results suggest that ß2-AR stimulation may improve the cardiac repair process by supporting an endogenous progenitor cell response but is not sufficient to improve the cardiac function.     

异翅负蝗配子发生过程中c-kit蛋白表达动态

应用免疫组织化学和生物统计分析相结合的方法,对异翅负蝗Atractomorp haheteroptera Bei Bienko配子发生过程中c-kit特异表达特点和动态进行研究。结果表明,(1)精子发生过程中,精原细胞、初级精母细胞、次级精母细胞和成熟精子中均有不同程度的c-kit蛋白表达,精巢末端还有较粗大的阳性颗粒分布;(2)卵子发生过程中,第1~6阶段卵母细胞中有不同程度的c-kit蛋白特异性表达,但随着卵黄发生的开始逐渐消失;(3)此外,滤泡细胞、输卵管和受精囊的腺细胞中有c-kit蛋白颗粒的存在;(4)从c-kit蛋白季节动态变化看,精子发生和卵子发生中的阳性表达都随着时间的延续出现下降的态势。因此,c-kit蛋白的特异性表达提示该蛋白参与配子发生过程的阶段性调控,具有重要的生理作用。     

糖尿病性膀胱病逼尿肌中SCFmRNA和SCF蛋白的表达

目的：检测DCP逼尿肌中SCF表达水平,探讨SCF基因表达与DCP关系及其发病机制。方法：按1：2病例对照研究,采用链脲佐菌素（STZ）及尿动力学检测成功建立DCP豚鼠20只为实验组,并以同质豚鼠40为对照组,应用RT-PCR和Western-blotting方法分别检测各组膀胱逼尿肌中SCF mRNA、SCF蛋白的表达。结果：DCP豚鼠组织中SCF mRNA表达与正常对照组比较无明显显著差异（P〉0.05）,DCP豚鼠组织中SCF蛋白表达明显低于正常对照组（P〈0.01）。结论：DCP组织中SCF蛋白表达减少与SCF基因翻译水平异常有关,因此高血糖环境下SCF基因表达异常可能是DCP的发病机制之一。     

Novel type of interstitial cell (Cajal-like) in human fallopian tube

We describe here--presumably for the first time--a Cajal-like type of tubal interstitial cells (t-ICC), resembling the archetypal enteric ICC. t-ICC were demonstrated in situ and in vitro on fresh preparations (tissue cryosections and primary cell cultures) using methylene-blue, crystal-violet, Janus-Green B or MitoTracker-Green FM Probe vital stainings. Also, t-ICC were identified in fixed specimens by light microscopy (methylene-blue, Giemsa, trichrome stainings, Gomori silver-impregnation) or transmission electron microscopy (TEM). The positive diagnosis of t-ICC was strengthened by immunohistochemistry (IHC; CD117/c-kit+ and other 14 antigens) and immunofluorescence (IF; CD117/c-kit+ and other 7 antigens). The spatial density of t-ICC (ampullar-segment cryosections) was 100-150 cells/mm2. Non-conventional light microscopy (NCLM) of Epon semithin-sections revealed a network-like distribution of t-ICC in lamina propria and smooth muscle meshwork. t-ICC appeared located beneath of epithelium, in a 10-15 microm thick 'belt', where 18+/-2% of cells were t-ICC. In the whole lamina propria, t-ICC were about 9%, and in muscularis approximately 7%. In toto, t-ICC represent ~8% of subepithelial cells, as counted by NCLM. In vitro, t-ICC were 9.9+/-0.9% of total cell population. TEM showed that the diagnostic 'gold standard' (Huizinga et al., 1997) is fulfilled by 'our' t-ICC. However, we suggest a 'platinum standard', adding a new defining criterion- characteristic cytoplasmic processes (number: 1-5; length: tens of microm; thickness: < or =0.5 microm; aspect: moniliform; branching: dichotomous; organization: network, labyrinthic-system). Quantitatively, the ultrastructural architecture of t-ICC is: nucleus, 23.6+/-3.2% of cell volume, with heterochromatin 49.1+/-3.8%; mitochondria, 4.8+/-1.7%; rough and smooth endoplasmic-reticulum (1.1+/-0.6%, 1.0+/-0.2%, respectively); caveolae, 3.4+/-0.5%. We found more caveolae on the surface of cell processes versus cell body, as confirmed by IF for caveolins. Occasionally, the so-called 'Ca2+-release units' (subplasmalemmal close associations of caveolae+endoplasmic reticulum+mitochondria) were detected in the dilations of cell processes. Electrophysiological single unit recordings of t-ICC in primary cultures indicated sustained spontaneous electrical activity (amplitude of membrane potentials: 57.26+/-6.56 mV). Besides the CD117/c-kit marker, t-ICC expressed variously CD34, caveolins 1&2, alpha-SMA, S-100, vimentin, nestin, desmin, NK-1. t-ICC were negative for: CD68, CD1a, CD62P, NSE, GFAP, chromogranin-A, PGP9.5, but IHC showed the possible existence of (neuro)endocrine cells in tubal interstitium. We call them 'JF cells'. In conclusion, the identification of t-ICC might open the door for understanding some tubal functions, e.g. pace-making/peristaltism, secretion (auto-, juxta- and/or paracrine), regulation of neurotransmission (nitrergic/purinergic) and intercellular signaling, via the very long processes. Furthermore, t-ICC might even be uncommitted bipotential progenitor cells.     

Cajal-type cells from human mammary gland stroma: phenotype characteristics in cell culture

We report here the in vitro isolation of Cajal-like interstitial cells from human inactive mammary-gland stroma. Primary cell cultures examined in phase-contrast microscopy or after vital methylene-blue staining revealed a cell population with characteristic morphological phenotype: fusiform, triangular or polygonal cell body and the corresponding (very) long, slender, moniliform cytoplasmic processes. Giemsa staining pointed out the typical knobbed aspect of cell prolongations. Immunofluorescence (IF) showed, like in situ immunohistochemistry, that Cajal-type cells in vitro (primary cultures), expressed c-kit/CD117 and vimentin. In conclusion, the images presented here reinforce our previous hypothesis that human mammary glands have a distinct population of Cajal-like cells in non-epithelial tissue compartments.     

Interstitial Cajal-like cells (ICLC) in atrial myocardium: ultrastructural and immunohistochemical characterization

We have previously reported (Hinescu & Popescu, 2005) the existence of interstitial Cajal-like cells (ICLC), by transmission electron microscopy, in human atrial myocardium. In the present study, ICLC were identified with non-conventional light microscopy (NCLM) on semi-thin sections stained with toluidine blue and immunohistochemistry (IHC) for CD117/c-kit, CD34, vimentin and other additional antigens for differential diagnosis. Quantitatively, on semi-thin sections, ICLC represent about 1-1.5% of the atrial myocardial volume (vs. approximately 45% working myocytes, approximately 2% endothelial cells, 3-4% for other interstitial cells, and the remaining percentage: extracellular matrix). Roughly, there is one ICLC for 8-10 working atrial myocytes in the intercellular space, beneath the epicardium, with a characteristic (pyriform, spindle or triangular) shape. These ICLC usually have 2-3 definitory processes, emerging from cell body, which usually embrace atrial myocytes (260 nm average distance plasmalemma/sarcolemma) or establish close contact with nerve fibers or capillaries (approximately 420 nm average distance to endothelial cells). Cell prolongations are characteristic: very thin (mean thickness = 0.15+/-0.1 microm), very long for a non-nervous cell (several tens of microm) and moniliform (uneven caliber). Stromal synapses between ICLC and other interstitial cells (macrophages) were found (e.g. in a multicontact type synapse, the average synaptic cleft was approximately 65 nm). Naturally, the usual cell organelles (mitochondria, smooth and rough endoplasmic reticulum, intermediate filaments) are relatively well developed. Caveolae were also visible on cell prolongations. No thick filaments were detected. IHC showed that ICLC were slightly and inconsistently positive for CD117/c-kit, variously co-expressed CD34 and EGF receptor, but appeared strongly positive for vimentin, along their prolongations. Some ICLC seemed positive for a-smooth muscle actin and tau protein, but were negative for nestin, desmin, CD13 and S-100. In conclusion, we provide further evidence of the existence of ICLC in human atrial myocardium, supporting the possible ICLC role in pacemaking, secretion (juxta- and/or paracrine), intercellular signaling (neurons and myocytes). For pathology, ICLC might as well be 'players' in arrhythmogenesis and atrial remodeling.     

Interstitial Cajal-like cells (ICLC) as steroid hormone sensors in human myometrium: immunocytochemical approach

Expression of estrogen (ER) and progesterone (PR) receptors was investigated in cultured human normal myometrial cells (non-pregnant uterus, fertile period). The ER and PR expression was studied by immunohistochemistry and immunofluorescence on either myocytes or interstitial Cajal-like cells (ICLC). Only those cells double immunostained for c-kit and steroid receptors were considered as ICLC. ER and/or PR immunoreactivity was localized in ICLC, primarily concentrated at the nucleus level, but it was also observed in the cell body (cytoplasm) and processes. Stronger immunopositive reaction in the ICLC nucleus for PR than for ER was noted. Under our experimental conditions, a clear positive repeatable reaction for steroid receptors could not be detected in myocytes. In conclusion, these data suggest that ICLC could be true hormonal 'sensors', possibly participating in the regulation of human myometrial contractions (via gap junctions with myocytes and/or by paracrine signaling).     

干细胞因子及其受体在红系分化中的作用

干细胞因子作为一种多能细胞因子,在临床上有较好的应用前景。随着CD34^ 细胞分离培养技术的完善,干细胞因子及其受体在红系增殖、分化中的重要作用正逐渐被发现并阐述清楚,对临床上相关疾病的治疗有重要的指导意义。本介绍了干系胞因子及其配体抑制红系祖细胞凋亡和促进红系祖细胞增殖和分化的功能,以及在临床上的应用前景。     

c-kit~+心脏干细胞在心脏再生修复中的应用

心血管疾病的流行是一个全球性的现象,在我国,其患病率也不断增长。目前尚无有效的治疗方法以解决心肌细胞损失这一关键问题,而干细胞移植很可能成为新的治疗方法。C-kit+心脏干细胞(cardiac stem/progenitor cell,CSCs)的发现证实了CSCs的存在,并为心脏的再生和修复治疗带来了新的曙光。C-kit+CSCs在心肌梗死、心力衰竭等心脏疾病中的作用得到了多项体内外实验及临床试验的证实,但由于目前尚缺乏直接的充分的证据证明内源性或外源性的c-kit+CSCs可分化为相当数量的成熟的具有功能的心肌细胞,其治疗机制尚存争议,同时,将其应用于临床仍面临多个问题。