Cell Line Construction And Function Exploration Of A Novel Type Of Kidney Interstitial Cell-Telocyte

Objectives:To isolate, identify and culture Telocyte (TC) in kidney. To unveil the distribution, morphological profiles, ultrastructure, phenotype of kidney TCs. To purify kidney TC and construct TC cell line. To identify whether TC is a kind of stem cell. To explore the influences of TC on the proliferation and apoptosis of tubular epithelial cell (TEC). To verify the paracrine effect of mesenchymal stem cell (MSC) on the proliferation and apoptosis of TEC. To detect whether TCs can influence the protection effect of MSC in kidney as a kind of stem cell supporting cell in vitro; To study the mechanisim involved in the communication between TCs and MSCs; To check the protection effect of MSCs conditioned by TCs in rat kidney ischemia-reperfusion model in vivo.Methods:We used transmission electron microscope (TEM), in situ immunohistochemistry on serial sections (IHC) and primary cell culture to determine the existence, distribution and phenotype of TCs in kidney. Furthermore, the ultrastructure of TC was revealed by TEM. Then immunofluorescence, vital cell staining and mitochondria labeling were used to identify TCs we found in kidney. TCs were isolated from kidney using primary cell culture. Then, we tried using mistiming adherence protocol, microdissection technique and passage to purify TCs. Cell-IQ was used to explore the growth rhythm of TCs and the morphological profiles which might be related to their potential role as a stem cell supporting cell were also uncovered. Then rat MSCs were cultured and identified using the previous methods. We used MillicellTM system to realize the cocultivations between TCs and TECs, MSCs and TECs, TCs and MSCs respectively, to explore the role of TCs in TECs proliferation and apoptosis, and to search the mechanisms involved in the communication between TC and MSC in vitro. Afterwards, we established the kidney ischemia-reperfusion model in rat. The protection effects of cell therapy through intravenous injection of MSCs conditioned by TCs in vivo were consequently evaluated by means of pathological and TUNEL scoring in rat kidney acute ischemia-reperfusion injury model. Results:There are TCs in human kidney cortex definitely. Kidney TCs are located in the peritubular and perivascular interstitial compartment. They often embrace the tubules and vessels tightly through their prolongations. Kidney TCs are characterized by their long, thin and moniliform prolongations (telopodes, Tps,), consisting of podoms and podomers. There are abundant mitochondria, vesicles and high-density particles in their cytoplasm. Kidney TCs are positive for CD117, CD34and Vimentin while negative for CD31, D2-40and Tryptase. These aspects distinguish TCs from other interstitial cells including fibroblasts and macrophages in kidney. In kidney medulla interstitium, however, no typical TC was observed in IHC and TEM images. But during primary cell culture of medulla samples, TCs emerged, accounting for only1%of total cell. Differentiation induction failed to yield positive result for TCs in vitro. Telocytes are not able to affect the proliferation and apoptosis of TECs, while MSCs can enhance the proliferation of TECs and prevent TECs from apoptosis as well. Conditioning MSCs using TCs can enlarge this effect of MSCs in vitro significantly, which indicates that TC might act as a kind of MSC supporting cell through paracrine pathway. Cell therapy through intravenous injection of MSCs conditioned by TCs can decrease the pathology scoring of the kidney suffering from ischemia-reperfusion injury in vivo. Meanwhile, kidney functions, resembled by SCr and BUN levels, were apparently improved.Conclusions:There are TCs corresponding to typical criteria in human kidney cortex. Kidney TCs possess abundant vesicles and high-density particles, which could be related to their paracrine effect. TCs can up regulate the proliferation promoting and apoptosis reducing effects of MSCs on kidney tubular epithelial cells by means of paracrine function. Cell therapy through intravenous injection of MSCs conditioned by TCs attenuates kidney ischemia-reperfusion injury significantly.