Influence of gene modification in biological behaviors and responses of mouse lung telocytes to inflammation

Abstract

Background Telocytes play key roles in maintenance of organ/tissue function and prevention of organ injury. However, there are great challenges to investigate telocytes functions using primary telocytes, due to the difficulties of isolation, identification, and stability. The present study aims at constructing continuous cell strain of mouse lung telocyte cell line with stable characters by gene modification and investigating biological behaviors and responses of gene-modified telocytes to inflammation. Methods Mouse primary lung telocytes were isolated and identified using immune-labeling markers and immunoelectron microscopy. Primary telocytes were transformed with Simian vacuolating virus 40 small and large T antigen (SV40). Biological characters, behaviors morphology, and proliferation of those gene-modified telocytes were defined and monitored dynamically for 50 generations, as compared with primary lung telocytes. Cell cycle of mouse primary lung telocytes or gene-modified telocytes was detected by flow cytometry. Results Gene modified telocytes of generations 5, 10, 30 and 50 were observed with telopodes and also showed CD34 and ckit positive. Multiple cellular morphology were also observed on telocyte cell-line under monitor of celliq and enhanced cell proliferation were showed. SV40 transduction was also reduced apoptosis and increased the ratio of S and G2 phases in telocyte cell-line. Conclusion We successfully constructed mouse lung telocyte cell-line which maintained the biological properties and behaviors as primary telocytes and could responses to inflammation induced by LPS. Thus, gene-modified lung telocytes, Telocyte Line, would provide a cell tool for researchers exploring the roles and applications of telocytes involved in physiological and pathological states in future. Electronic supplementary material The online version of this article (10.1186/s12967-019-1870-y) contains supplementary material, which is available to authorized users.

Telocytes inhibited inflammatory factor expression and enhanced cell migration in LPS-induced skin wound healing models in vitro and in vivo

“…To maintain microenvironmental haemostasis, complex communication with adjacent or distant cells is required for a special location within connecting cells [10]. The types of cells connected to TCs include fibroblasts, epithelia, myocytes, and other organ cells [6].…”

“…Mouse TCs were kindly provided by Dr. Dongli Song (Zhongshan Hospital Institute for Clinical Science, Fudan University, Shanghai, China). This TCs were isolated from mouse and transfected with SV40 large and small T antigen, and identified with telopodes (Tps) and the expression of ckit, CD34, vimentin and PDGFRα from generation 5 to generation 50 [10]. Human dermal endothelial cells (HDMECs), human immortalized keratinocyte (HaCaT) cell line, and L929 mouse fibroblast cell line were purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China), and they were cultivated in the recommended media and conditions.…”

“…Recently, studies have demonstrated that TCs exist in skin tissues according to focused ion beam scanning electron microscopy (FIB-SEM) tomography and with the establishment of the 3D reconstruction of dermal TCs [9]. Song et al recently established a mouse TC cell line (TCs) and demonstrated the maintenance of behavioural morphology and biological characteristics for 50 generations, which provided further patterns for the TC study [10]. However, whether TCs can promote skin wound healing as well as the mechanisms involved in this process remain unclear.…”

A Two-Step Immunomagnetic Microbead-Based Method for the Isolation of Human Primary Skin Telocytes/CD34+ Stromal Cells

“…Of note, over the past few years, several attempts have been made to isolate TCs from various organs such as the uterus [ 39 , 44 , 46 ], lung [ 45 ], and heart [ 41 , 42 , 43 ]. Nevertheless, it is important to consider that all these studies discerned TCs only on the basis of their different adhesion time on the plate, placing into culture the cells remaining in suspension after fibroblast attachment [ 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 ] and assessing their immunophenotype only subsequently [ 40 , 44 , 46 ]. In this context, our cell purification approach, which consists in the selection of TCs immediately after tissue processing and according to their immunophenotypic profile, may represent an innovative, reliable, and reproducible method to obtain a pure TC population suitable for in vitro analyses and other applications.…”

“…Despite the growing belief that TCs may exert important roles in both physiologic and pathologic conditions, at present, the majority of studies investigating these cells are mainly descriptive and focus only on their morphology, with a consequent paucity of functional data. To gain relevant insight into the possible functions of TCs, in vitro analyses are clearly required, but currently, the protocols for TC isolation are only at early stages and not fully standardized [ 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 ]. The present study was, therefore, undertaken to develop, for the first time, a methodology for the selective in vitro isolation of human primary TCs/CD34+ stromal cells consisting in a two-step immunomagnetic microbead-based separation of these cells from two other stromal cell populations, namely ECs and fibroblasts.…”