Subaqueous acoustic pressure system based one day heterotypic pseudo-islet spheroid formation with adipose derived stem cells for graft survival-related function enhancement
To overcome Type 1 diabetes mellitus (T1DM), which can cause hyperglycemia due to diminished insulin secretion of β-cell function, islet transplantation has been developed with various strategies including pseudo-islet. However, conventional pseudo-islet formation techniques combining with other cells depend on natural cellular aggregation, which requires at least 5 days to form and even show segregation of distinct cell types, leading to diminished cell viability and function. Herein, we applied a subaqueous free-standing 3D cell culture (FS) device, which can reduce the spheroid formation time by trapped cell in nodes of acoustic standing wave. Briefly, Culturing with adipose-derived stem cells (ADSCs) to form heterotypic pseudo-islet (Hislet) in FS device dramatically reduced formation time less than one day. Hislet demonstrated enhancement of cell viability than conventional pseudo-islet formation method. Additionally, ADSCs combined Hislet proved strong secretion of various paracrine factors. Also results showed significantly increased angiogenesis effect and immunomodulation effect for various type of immune cells in Hislet compared to islet, which can enhance transplantation survival. Furthermore, Hislet validated glucose-regulating capacity and enhanced angiogenesis effect in vivo T1DM model. Throughout this study, we propose a novel strategy for forming Hislet that can overcome the limitations of conventional Islet and pseudo-islet for T1DM.
To further evaluate the long-term glucose-regulating capacity and therapeutic potential of Hislet for T1DM, in vivo transplantation studies were conducted. After Specifically, Hislets (Equivalent to 400 IEQ) were transplanted under the kidney capsule of BALB/c nude mice (Fig. 6A). Non-fasting blood glucose (NBG) levels were monitored for 21 days post-transplantation. Islets were used as a control group, while the experimental group received Hislets. Both the islet and Hislet groups successfully maintained normoglycemic levels throughout the observation period (Fig. 6B), indicating that Hislet retained functional glucose-regulatory ability in vivo, comparable to that of Islet. To confirm that glycemic control was mediated by the transplanted grafts rather than endogenous insulin recovery, the grafts were surgically removed on day 21. Following graft removal, both groups exhibited a rapid return to hyperglycemia, demonstrating that blood glucose regulation was directly attributable to the transplanted grafts (Fig. 6B, Day 21). To further assess the glucose responsiveness of the grafts under physiological conditions, an intraperitoneal glucose tolerance test (IPGTT) was performed on day 20 post-transplantation. Hislet-transplanted mice exhibited a less maximum blood glucose level during IPGTT and lower area under the curve (AUC) compared to mice transplanted with na\u00EFve islet (Fig. 6C\u2013D) supporting the notion that Hislet exhibit enhanced glucose responsiveness and insulin secretion capacity. Immunofluorescence staining of graft-bearing kidneys harvested on day 21 revealed comparable insulin-positive cell masses in both groups (Fig. 6E). However, CD31 expression, a marker for vascular endothelial cells [91] was more prominently observed in the Hislet group, suggesting enhanced angiogenesis at the graft site.Ki Dong Park reports financial support was provided by Republic of Korea, Ministry of Trade, Industry & Energy. Jee-Heon Jeong reports financial support was provided by Republic of Korea, Ministry of Science and ICT. Hyun-Ji park reports financial support was provided by Republic of Korea, Ministry of Science and ICT and Ministry of Trade, Industry & Energy. Dong Yun Lee reports financial support was provided by Republic of Korea, Ministry of Science and ICT, the Ministry of Health & Welfare. Suk Ho Bhang reports financial support was provided by Republic of Korea, Ministry of Science and ICT, the Ministry of Health & Welfare and Ministry of Trade, Industry & Energy. Dong Yun Lee reports a relationship with Elixir Pharmatech Inc. That includes: employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.This work was supported by the National Research Foundation (NRF) grants funded by the Ministry of Science and ICT (Grant No. RS-2023-00272815 to J-H.J. RS-2024-00411474 to H-J.P) and the Korean Fund for Regenerative Medicine (KFRM) grant funded by the Korea government (the Ministry of Science and ICT, the Ministry of Health & Welfare) (No. 21A0102L1-13 to S.H.B. and KFRM24A0105L1 to D.Y.L. Republic of Koea). This work also supported by the Alchemist Project of the Korea Evaluation Institute of Industrial Technology (KEIT 20018560, NTIS 2410005252 to S.H.B. H-J.P, and K.D.P), the Ministry of Trade, Industry & Energy, Republic of Korea.
