Sustained Effect Of Hyperglycemia And Calcitriol On Native Mechanical Tendon Properties

Objective: Diabetes mellitus is a complex metabolic disorder which can cause detrimental changes to different systems.Previous studies have revealed that the diabetic state is associated with osteoporosis[1],nephropathy[2],Cardiovascular Complications[3],and retinal pathology[4].But the impact of hyperglycemia on the tendon has only recently been examined.A few studies indicated diabetes mellitues(DM)having negative impacts on the mechanical properties of the tendons.At present the study of diabetes span ranged from several days to weeks,mainly focused on the Achilles tendon,it shows that the mechanical property of the tendon was reduced,such as reduced mean load to-failure and stiffness of the tendon-bone complex[5],reduced Young's modulus[6],lower stiffness and peak load in the injured diabetic tendons[7].As we all know,calcium is a recognized second messenger implicated in insulin secretion.Vitamin D(1,25-dihydroxycholecalciferol,Calcitriol)plays an important role in calcium metabolism.Previous study has determined that Calcitriol improves diabetic nephropathy in rats by inhibiting renin and relieving oxidative stress,treating diabetic cardiomyopathy[11],suppressing inflammation,lowering albuminuria[8-10],recovers bone mineral density[12] and increase insulin sensitivity[13].Unfortunately,there was no study designing to determine if Calcitriol could restore biomechanical properties of patellar tendon in diabetic rats.Therefore,the objective of our study was to determine the native mechanical properties of the patellar tendon and the protection of calcitriol between healthy control and hyperglycemic rats 9 weeks following induction of hyperglycemia with intraperitoneal(IP)injection of streptozotocin(STZ).This study was tested using a randomized controlled model of streptozotocin-induced diabetes in rats.Methods: Sixty-five male Sprague–Dawley rats were divided randomly into a model group(n =50)and a control group(n =15).Then the model group was injected with a maximum of three 30 mg/kg doses of STZ dissolved in a citrate buffer every 3 days until hyperglycemia was present.According to the protocol to induce diabetes described by Thomas.Control animals received an IP injection of citrate buffer solution only [14].Hyperglycemia was confirmed with a fasting blood glucose level of ?11.1mmol/L with a glucometer.The rate of successful model establishment was about 80.3% which has been known from our pre-experimental results.Forty rats which confirmed the hyperglycemic rats were divided into a hyperglycemia group(n =20)and a calcitriol group(n =20).The calcitriol group was injected with a dose of 1mg/kg of the animal's weight every day for nine weeks while other animals received an IP injection of oil only.All the animals remained in normal cage activity and had free access to water and chow over 9-week period.During the 9 weeks,fasting blood glucose levels were recorded weekly to detect the change of glycemia.Animals were excluded if fasting blood glucose levels were less than 11.1mmol/L every measurements.The three groups were then sacrificed 9 weeks following hyperglycemia induction.(1)measure the length,width,thinness of the tendon by digital micrometer;(2)stained with hematoxylin and Eosin to assess collagen organization and tendon-patella complex;(3)The complex were designated for mechanical testing(stiffness and creep);(4)measure the level of 1,25-dihydroxy –cholecalciferol.Results:1 The hyperglycemia and calcitriol group had a significantly higher fasting blood glucose level compared to controls all the time.The level of calcitriol group was slightly lower than the hyperglycemia group,but these alterations were not statistically significant.2 There were significant differences in the level of 1,25-dihydroxycholecalciferol among three groups.The level of hyperglycemia group was lower than the control group,while the calcitriol group has improved.3 The gross and histological findings were similar to the previous observation.The tendons of the diabetic rats were shrinking,fragile and yellowish,compared with the robust,healthy tissue in control animals.4 There were significant differences in length and width between Hyperglycemia and Control groups.However,the measurements was underpowered to detect statistically significant differences in Hyperglycemia and Calcitriol groups.Despite all that.Calcitriol could better tendon degeneration which could demonstrate from the greater length and width compared with the hyperglycemia groups.5 In contrast to the tight,parallel,bundled appearance in the control tendons,there were plenty of small tears in the diabetic tendons,what is more,the collagen fiber were crimped and irregular with decreased cell density.However,the Calcitriol group had similar appearance with the control group.6 the mean stiffness of the diabetic patellar tendons was decreased compared to control tendons.A significant difference in stiffness of the tendon was also detected between Calcitriol and control groups,meanwhile the Hyperglycemia and Calcitriol groups also demonstrated a significantly difference.The tendons of the diabetic group exhibited lower creep property compared to the control tendons loaded to either 5N or 10 N.The differences were statistically significant.However,the creep date of Calcitriol group was similar to the diabetic group,the effect of Calcitriol on the creep property of diabetic tendon was doubtful.The maximum tensile load is 200 N,10 of 15 tendon of diabetes group ruptured in experimental process,and in the normal control group only 3 tenden teared,in calcitriol group the number of rupture tendon is 5,the rest to the end are intact.Obviously the maximum tensile load of diabetes group decreased.These reduced biomechanical properties may lead to the easy tendon rupture.Conclusions: In conclusion,nine weeks of hyperglycemia led degeneration in tendon mechanical properties,such as a significantly reduced creep property and stiffness of the patellar tendon compared to control specimens,as well as the crimped and irregular collagen fiber with decreased cell density.It is worth noting that a significant difference in stiffness of the tendon was detected between hyperglycemia and calcitriol groups,in addition,we found the improved mechanical property of the Calcitriol group tendon with the beneficial effects on blood glucose,weight and water consumption in diabetic rats.These findings provided a new proposition to make a preventive strategy for diabetic tendon degeneration.