JOURNAL OF MILITflRV PHflRMflCO-MCDICINC N°7-2014
RESEARCH ON THE CHANGES OF BLOOD GLUCOSE LEVELS, INSULIN CONCENTRATIONS AND AMYLASE ACTIVITIES IN
PORCINE PANCREATIC TRANSPLANTS
Vu Quang Hop*; Pham Van Tran**
SUMMARY
The aim of this research is to determine the changes of blood glucose level, insulin concentration and amylase activity on 30 pigs after pancreatic transplantation Pigs were transplanted with total pancreatic allograph. Bloods for biochemical test were obtained before and after surgery at the time 3, 24, 48, 72 and 96 hours. Results: Blood glucose level before surgery was 4.93 ± 2.9 mmol/L, after surgery at the time 3. 24, 48, 72. 96h. these parameters were respectively 8 1 ± 5.5; 4.97 ± 2.8; 4.96 ± 2.2; 5.1 ±2 1; 4.9 + 2.0 mmol/L. Blood insulin level before surgery was 4.0 ± 2.6 plU/mL, after surgery at the time 3, 24, 48, 72, 96h, these parameters were respectively 5.5 ± 2.2; 36 ± 2.4; 3.1 ±2.0; 2.9 ± 1.7; 23 ± 1.1 plU/mL Blood amylase activity before surgery was 1,295 ± 496 U/L, after surgery at the time 3, 24. 48, 72, 96h, these parameters were respectively 1.569 ± 923; 1,365 ± 504; 1,442 ± 575; 1.394 ± 574; 1558 ± 502 U/L. Conclusion: In the pigs transplanted with total pancreatic allograph blood glucose level and amylase activity increased highly at 3h after surgery then gradually decreased but always higher than that before transplantation while blood insulin level increased at the 3h after transplantation (p < 0.05). The next time after transplantation, blood insulin levels tend to decrease gradually lower than these levels before transplantation
* Key words: Pancreatic transplantation; Glucose; Insulin; Amylase; Pig.
INTRODUCTION disease, stroke, kidney failure, foot ulcers and damage to the eyes.
Diabetes mellitus (DIVI) also known as „ , ^ „ , „ „ . . „ .- , , « „ „ Globally, as of 2013, an estimated 382 simply diabetes, is a group of metabolic ^^^.^^ p^^p,^ ^^^^ ^i^^jetes worldwide, diseases in which there are high blood ^/jth type 2 diabetes making up about 90%
sugar levels over a prolonged period. This of the cases This is equivalent to 8.3% of high blood sugar produces the symptoms the adult population, with equal rates in of frequent urination, increased thirst, both women and men. In the world and increased hunger. Untreated diabetes
can cause many complications. Acute
2012 and 2013, diabetes resulted in 1.5 to 5.1 million deaths per year, making it the 8^
leading cause of death. Diabetes overall at complications include diabetic ketoacidosis ,533, ^^ubles the risk of death. The number of and nonketotic hyperosmolar coma. Serious people with diabetes is expected to rise to long-term complications include heart 592 million by 2035 [1,2],
• 69 Institute of Hochiminh Mausoleum
" 103 Hospital
Corresponding author: Phan Van Tran ([email protected])
67
JOURNRl OF MIUTRRV PHRRmnCO-MCDICINC N°7-20U However, most current therapies do not cure the diabetes. In this context, scientists in the worid are constantly trying to find new therapies to treat diabetes.
Actually, one of the most advanced methods is pancreatic transplantation. Whole pancreas transplantation was first performed in 1966 at the University of Minnesota (USA).
Cun-ently, each year there are about 1,300 people vnth type 1 diabetes whose pancreas transplanted [3].
In Vietnam, the scientists have conducted transplant and has gained a lot of success.
However, pancreas transplantation is still very new and no studies have been done.
Before proceeding human organ transplant, we may undergo experimental transplantation on animals. It was also agreed that a new surgical methods should be studied in all aspects and the surgeons should be well trained on expenmental techniques.
Pancreas transplant involved biological changes happening inside the body wrtiich received the particular biochemical changes. We conducted this subject with the aim of identifying some blood biochemical parameters in pigs before and after pancreas transplantation in order to provide a basis for successful research on human pancreas transplant.
ANIMALS AND METHODS 1. Animals.
We conducted 30 pairs of pig 3 - 4 months old, weighing from 45 - 60 kg, healthy for pancreas transplant (60 pigs).
Pigs were fed by a special 3 days diet in strict hygienic conditions. Each experimental surgical procedure needs h«o pigs which are in the same mother to ensure maximum harmony immune factors; 1 for pancreas donor and 1 for pancreas recipient.
2. Methods.
The receiving pigs were incised in white line abdomen between the upper and lower umbilical point to remove the entire pancreas, then the donor's pancreas was transplanted into pelvic pits of recipient.
Receiving pigs were kept for 4 days after surgery. Blood was taken (3 ml) with heparin as anticoagulant substance for biochemical test before and after surgery at the 3, 24, 48, 72 and 96 hours Bloods were centrifuged for obtaining plasma. Biochemical panameteis were performed by Au640 system using kits of Beckman Coulter in Department of Biochemistry, 103 Military Hospital.
The collected data were calculated by the method of biostatistics using SPSS software V16.
RESULTS
1, Results of pancreas transplantation in pigs.
Our number of transplanted pigs is 30.
During and post-operation, the survival time of pigs shown as below.
* The survival time of pigs:
Death dunng operation: 2 pigs; living after 24h; 28 pigs; living after 48h:20 pigs; living after 72h; 15 pigs; living after 96h: 4 pigs.
Two pigs died during operation. Post operation, survival time were different and only 4 pigs were survival after 96h.
* Classification of surgical complications (n - 28):
Embolism; 13 pigs (46,4%); hemorrhage;
4 pigs (14,3%); infection; 9 pigs (32,1%);
pancreatitis; 2 pigs (7,2%).
We can see the pancreas is purple with lesion; it may be also accompanied by intestine necrosis.
68
JOURNRl OF MIUTflRV PHRRMRCO-MEDICINC N ° 7 - Z a i 4 2. Results on biochemical parameters before and post-operation,
* Stood glucose levels:
Table 1: Glucose levels (mmol/L).
x
SD Min Max P
BEFORE OPERATION
(n-30)(1) 4.93
29 1 1 16
POST-OPERATION 3h
(n = 28)(2) 6.1 5.5 2.9 21.2
24h (n = 28) (3)
4 97 2.8 2.3 17
48h (n = 20) (4)
4,96 2.2 1.4 12
72h (n = 15)(6)
5 1 2 1 3 2 11
96h (n =4) (6)
49 2.0 2.2 68 Pi.2 < 0 05; pi.3 > 0.05, Pi.* > 0.05; pi.s > 0 05: piji > 0 05
Before operation, blood glucose levels were 4.93 ± 2.9 mmol/L, which increased highly at 3h post-operation (p < 0.05), then decreased with the time to normal value
* Blood insulin levels:
Table 2; Blood insulin levels (plU/mL).
x
SD Mm Max P
BEFORE OPERATION
(n = 30)m 40 26 0.9 9.4
POST-OPERATION 3h
(n = 28)r2J 5.5 2.2 2.3 99
24h (n = 28) (3)
36 24 08 9.1
48h (n = 20) (4)
3.1 2.0 09 81
72h (n = 15) (5)
2.9 1-7 0,5 6-7
96h (n = 4) (Sj
2.3 1 1 1.1 37 Pi-2<0.05; p,.3<0.05: pi^<0 05: Pi-5<0 05. pi.a,<0.05
Before operation, blood insulin level was 4 ± 2.6 (plU/mL), which inaeased at 3h post-operation (p < 0 05). After the operation, it tends to decrease lower than this parameter before graft.
* Blood amylase activity:
Table 2: Blood amylase activity (U/L),
X SD Min Max P
BEFORE OPERATION (n = 30)ffl
1295 496 599 2369
POST-OPERATION 311
(n = 28) (2) 1569 923 409 4716
24h (n = 28) (3)
1365 504 684 2549
48h (n = 20) (4)
1442 575 693 2572
72h (n = 15) (5)
1394 574 789 2645
96h (n = 4) (6)
1558 502 1110 2160 P M < 0.05, Pi.3 < 0,05: Pi.4 < 0 05; pi.s < 0,05, p i ^ < 0 05
Before operation, blood amylase activity was 1.295.2 ± 496.6 (U/L) After the operation,
JOUANAl OF MillTRRV PHRRMRCO-MeDICINC N°7-2014 DISCUSSION
1. Successful pancreas transplantation in pigs.
We choose the pair of pigs of the same mother to reduce maximum the difference in the immune system related to pancreatic allograph rejection. According to the previous studies on pig transplants showed that pig anatomy and physiology similar are to humans. Thus, from this study, the results can be applied to the research on the human pancreas transplant.
For such studies, the endocrine pancreas function of recipient should be abolished before the transplantation. The best way to achieve this is to perform a total pancreatectomy in the recipient sometimes before transplantation.
A number of findings from the preceding experiments were applied to these transplantation experiments. Since it had been found that functioning exocrine pancreatic tissue could still be present after dissection of pancreas from the duodenum, a number of pancreas function tests had to be performed after total pancreatectomy to demonstrate the abolition of both exocrine and endocrine functions.
Furthermore, the pancreatitis v^ich invariably accompanied with duct ligation led to a rise in the hematocrit value with sludging of erythrocytes, increasing the risk of intravascular coagulation.
This made it advisable to supply fluids abundantly after pancrea transplantations.
Lastly, in connection with the autotnansplantatons it was found that hypoglycemia could develop after operation, and we therefore administered glucose intravenously in the postoperative period The average weight of pigs is 50 - 60 kg, equivalent to the weight of the people, so surgeons can standardize operations for practicing the technique before work on human later
The dominant post-transplant complications are bleeding and venous thrombosis. The vascular embolism reduced the circulation in pancreas making necrosis. Two pigs died during the operation or within the first 24 hours, one due to an severe bleeding and one from severe post-operative hypoglycemia despite the administration of more than standard amount of glucose
2. Changes of blood glucose, insulin levels and amylase activity.
* Blood glucose levels:
Following table 1, before operation, blood glucose level was 4.93 ^ 2.9 mmol/L, corresponding to that in human blood.
This value is equivalent to the people.
Blood glucose levels increased during and post-operation, particularly after 3 hours. This is explained by the pancreas removed, so insulin doesn't seaete causing hyperglycemia. The other researchers also did the same experiments in dog, after cutting the pancreas, blood glucose increased and dogs became diabetic 11, 4], At 3h post-operation, pancreas did not activate. The researchers showed that, the transplanted pancreas start secrete insulin after 12h post operation [5, 6], 70
JOURNfll OF MIUTRRV PHRRMACO-MCDICINC N°7-2ai4 The next time after transplantation, the
pancreas had returned to active secretion of pancreatic hormones including insulin into the bloodstream. Blood glucose had returned to the time before surgery but then Increase or decrease unstable. This is also consistent with the clinical and physiological pancreas transplant as well and have not recovered elements of glucose before, during and after surgery.
* Blood insulin levels:
Table 2 showed that before operation, blood insulin was 4.0 ± 2.6 Ul/ml, then decreased gradually in the next moment In the body, insulin was secreted by the beta cells of the pancreatic Islets which have more effective at reducing blood glucose. Chronic pancreatitis may increase or decrease insulin. Insulin reduced in the case of damage islet beta cells or removed pancreas. The insulin value is much more important for the evaluation of endocrine pancreas function. When insulin levels in the blood are normal or increased blood glucose but can still Increase due to the body's cells are no longer sensitive to insulin, insulin decreased in our study reflected by pancreatic lesions, pancreatic islet beta cells reduce insulin secretion.
Blood glucose and insulin levels reflected damage evolution of islet cells. Its rise and decline can predict the results of pancreatic transplantation.
* Blood amylase activity:
The serum amylase level varied widely in all of groups. Tablet 3, before operation, blood amylase activity was 1,295 ± 496 U/L.
According to previous studies, it was 1,150 + 103 U/L [6]. The results in table 3, amylase activity tends to increase after transplantation, especially at 3h after transplantation. Then, it was maintained a high level in the next moment. Elevated blood amylase activity is considered as a sign of acute pancreatitis. After transplant, pancreas damaged by the inflammatory response in place, in addition due to mechanical trauma during surgery. Elevated blood amylase activity could be similar in pancreatic duct obstruction. Amylase blood started to be increase from 2 hours to 12 hours after acute pancreatitis and returned to normal within 3 - 5 days [3]. In the case of transplantation of pancreas, amylase more slowly return to normal range after 1 - 2 weeks [7]. This is also consistent with the results of our research. In addition, many scientific reports showed that in the pig pancreas transplant, anti-rejection drug as well solumedrol could make amylase activity higher than normal.
CONCLUSION
The results led us to conclude that in the pigs transplanted with total pancreatic allograph, blood glucose level and amylase activity increased highly at 3h after surgery then gradually decreased but always higher than which before transplantation while blood insulin level than increased at the 3h after transplantation (p < 0.05). The next time after transplantation, blood insulin levels tend to decrease gradually lower than these levels before transplantation.
71
JOURNAL OF MrUTARV PHflRMnCO-MCDICINC N<>7-2014
R E F E R E N C E S
1 Bonora £ Protection of pancreatic beta- cells is it feasible? Nutr Metab Cardiovasc Dis.
2008, 18(1), pp.74-83.
2. Chaib E. GalvSo FH. Total pancreatectomy:
porcine model for inducing diabetes - anatomical assessment and surgical aspects, Eur Surg Res.
2011,46(1), pp 52-55,
3. George k. Kyriakides. Porcine pancreatic transplants. Department of Surgery and Medicine Minesota. 1975.
4 Han DJ, Sutherland DE._ Pancreas transplantation. Gut Liver 2010, 4 (4), p.450- 465. Epub.
5. Joana Ferrer Pig pancreas anatomy.
Implications for pancreas procurement, preservation, and islet isolation T r a n s p l a n t a t i o n . 2008, pp.1503-1510.
6. Kelly WD. Lillehei RC. Allotransplantation of the pancreas and duodenum along with the kidney in diabetic nephropathy. Surgery. 1967, 61 (6), pp 827-837.
7. Lillehei RC. Pancreatico-duodenal allotransplantation: experimental and clinical experience. 1970, 172 (3), pp.405-436.
7 2
