ABSTRACT
BACKGROUND: Based on previous technique prepared for encapsulating living cells with alginate-polysine- alginate (APA) microcapsules, it has been confirmed that microencapsulated chromaffin cells have good analgesic effects. The immunoisolated effects of such microcapsule materials need to be evaluated. OBJECTIVE: This study aimed to investigate the immunological rejections of APA microencapsulated chromaffin cells transplanted into rat anterior chamber of eyes and tendon of feet, and to evaluate the immunoisolated effect of microencapsulation.DESIGN: A randomized controlled animal experiment. SETTING: Department of Anesthesiology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: Forty-eight female SD rats, with the age of 3 months, were provided by the Laboratory Animal Center, Tongji Medical College, Huazhong University of Science and Technology. The protocol was carried out in accordance with ethical guidelines for the use and care of animals. Alginate and polylysine used in the experiment were the products of Sigma Company, USA. Microcapsule generator was gifted by Germany. METHODS: This study was performed at the Department of Anesthesiology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology from September 2002 to September 2003. Suprarenal medulla was taken from 6 healthy adult cadavers of brain death. After isolated, digested and cultured, suprarenal medulla was prepared into chromaffin cell suspension. Written informed consents were obtained from the family members of donors, and the protocol was given approval by the Ethics Committee of the hospital. Empty microcapsules and microencapsulated cells were prepared by APA. The 48 rats were randomly divided into the human chromaffin cell (HCC) group, the empty microcapsule group and the microencapsulated HCC (ME-HCC) group. In each group, there were two transplanted regions of anterior chamber of eyes and tendon of feet, with 8 rats used for each region. Each rat in the HCC group was perfused 2×1010 L-1 cell suspension into the anterior chamber of eyes and tendon of feet. Those in the empty microcapsule group and the ME-HCC group were perfused 100 empty capsules and ME-HCCs (100 microcapsules, 400-500 HCCs per microcapsule) into the same regions, respectively. MAIN OUTCOME MEASURES: On day 7 after transplantation, serum interleukin (IL)-2 level was determined by ELISA. Serum IgG and IgM levels were determined with a laser turbidimeter. On day 28 after transplantation, rat right eyeball and left feet were harvested, routinely sliced and stained by haematoxylin-eosin (HE). Histo-morphological structure was observed under a 40×light microscope. RESULTS: Forty-eight rats were included in the final analysis. Serum IL-2, IgG and IgM levels were significantly lower in the empty microcapsule group and ME-HCC group than in the HCC group (t=8.544-21.64, P < 0.01). A lot of lymphocyte and neutrophile infiltration could be found in the anterior chamber of eyes and tendon of feet of rats in the HCC group, but a little seen in that of the empty microcapsule group and ME-HCC group. CONCLUSION: APA microencapsulation has an effective immunoisolated effect on immunological rejection due to its good biocompatibility and mechanical stability.
ABSTRACT
Objective To compare the catecholamine (CA ) and M enkephalin(M ENK) release from human chromaffin cells (HCC) and microencapsulated HCC (ME HCC ) and investigate the effects of microencapsulation on the growth and activity of chromaffin cells Methods Adrenal glands were taken from brain death healthy adults The chromaffin cells were isolated and primarily cultured in vitro During culture the chromaffin cells were HE stained and assessed by labelling the cells with tyrosine hydroxylase monoclonal antibody The percentage of tyrosine positive cells were counted under microscope (1) The chromaffin cells were microencapsulated with 2% alginate and cultured in vitro (experimental group) HCC which were not microencapsulated were used as control The culture media of both groups were replaced every 48h and collected and stored under -20℃ for determination of CA and M ENK concentration (2) On the 6th day of culture, nicotine was added to HCC and MC HCC suspension After 30 min incubation, the suspension was centrifuged and the supernate collected and stored under -20 ℃ for determinatoin of CA and M ENK concentration with radioimmunoassay Results (1) ME HCC grew fairly well in vitro in the culture medium and was morphologically similar to HCC (2) There was no significant difference in CA and M ENK concentration in HCC and ME HCC culture media (3) CA and M ENK concentrations in supernate were increased by nicotine stimulation and there was no difference in the CA and M ENK concentration in the supernate between the two groups Conclusions Alginate and microencapsulation technique are not harmful to HCC, ME HCC has fairly good activity and release function and can be effectively used for transplantation
ABSTRACT
Objective To compare the survival of isolated chromaffin cells with that of adrenal medullary pieces implanted into the subarachnoid space .Methods Forty male Sprague Dawley rats, 180 220g, were randomly allocated to be implanted into the subarachnoid space with the homologous adrenal medullary pieces washed with PBS(group A,n=20) or the isolated chromaffin cells 5?107 in 10?l suspension (group B,n=20), respectively, after the implants were in vitro cultured in three days . The thermal threshold was determined before ,5 and 10 weeks after the transplantation. Five or ten weeks after the transplantation, 6 rats of either group were randomly selected to obtain the adrenal medullary grafts from the subarachnoid space or the sediments of cerebrospinal fluid ,observed with an electron microscopy.Results Five weeks after transplantation, the thermal threshold in both groups increased markedly,but without significant difference between them ,and there were intact chromaffin cells in the sections of both groups. Ten weeks after the operation, the thermal threshold in group A decreased to basaline, and was significantly lower than in group B; no intact chromaffin cells were found ,with the infiltration of a large number lymphocytes in the sections of group A , and in the sections of group B, the intact chromaffin cells were found with the infiltration of lymphocytes in lower amount and density. Conclusions The isolated chromaffin cells implantation is superior to the adrenal medulla pieces implantation for analgesia.
ABSTRACT
Objective To investigate the immunoisolation effect of xenografts of microencapsulated human chromaffin cells (HCCs) .Methods HCCs were microencapsulated with APA microcapsules. Forty-eight SD rats were randomly divided into 3 groups ( n = 16 each) . HCCs (group I ) , empty APA microcapsules (group II ) and microencapsulated HCCs (group III ) were implanted into the anterior chamber of the eyes and the sole of the feet. Seven days after transplantation blood samples were collected for determination of serum IL-2, IgG and IgM concentration. The right eye-balls and left feet were obtained for microscopic examination. Results The serum IL-2, IgG and IgM concentrations were significantly lower in group II and group III than in group I ( P