The Therapeutic Effection And Mechanism Of GM-CSF On Wound Healing With Type2Diabetic Rats

Objective: Diabetic patients with post-traumatic wound healing is not easy for thecommon clinical problem, especially in patients with type2diabetes. This experimentby using a rat model of type2diabetes, to investigate the effect of exogenous GM-CSFon type2diabetic rat wound healing effect and mechanism.Methods: Take February age SPF female rats70, body weight (200+15) g, inhigh-fat feed feeding for6weeks after the start of environment modeling, modelingfasting12hours before, but the water. STZ prepared with new0.1mmol/L citric acidbuffer solution (PH=4.2) dilution, rats after intraperitoneal injection of STZ weighingdisposable35mg/kg,48hours after the blood glucose, blood glucose monitoring.Selection of modeling success of55rats in10%chloral hydrate (300mg/kg)intraperitoneally anesthesia back after shaving, in the back by using the midline thighboth sides die diameter1.5cm round wound, wound area of about1.77cm L. Will theskin along the line cut, deep to the deep fascia. Random modeling success of40ratswere divided into2groups: control group for diabetes (DM) wound in20only, theexperimental group for diabetic wound+GM-CSF20(GM-CSF0.5ug/kg woundinjection). Another15rats were divided into5groups, each group of3. Group A: DMgroup; group B: DM+intraperitoneal injection of saline group; group C: DM+intraperitoneal injection of GM-CSF; D: DM+local GM-CSF; group E: DM+localsaline group, rats cage rearing. The eye of rats were observed after third,5,8,12,18daysof wound healing, at the same time every day to rats weighing, in the tail vein bloodglucose monitoring value. Records of each rat body weight, blood glucose changes andphysical state changes. Use of transparent paper along the outer edge of third, depictingthe wound healing of5,8,12,18days of the wound area, calculating the wound healingrate. Along the healing of the wound edge in excess of approximately1cm cut wound tissues were observed under optical microscope, production various types of cells, deeptissue wound healing. Sampling line transmission electron microscopic observation offibroblast morphology, immunohistochemical observation of PCNA, BCL-2positivecells.Results: At each time point, GM-CSF injection drugs in experimental group isobviously higher than that of control group, the average area of healing. During thehealing period, the experimental group averaged19.50+/-1.29days, the control groupwas22+/-0.82days. HE staining: experimental group third days in the presence of alarge number of wound exudate and the influx of neutrophils and other inflammatorycells, cell type, quantity; controls cell number is small, the organizational structure isnot clear. The experimental group fifth days of organization structure is clear, start thepresence of small amounts of granulation tissue and glandular structures; the controlgroup with massive infiltration of cells, cells arranged in confusion. The experimentalgroup8-12day visible amount of fibroblasts, collagen bundles and skin structure,organizational stratification of healing, appeared on the cortex and connective tissuelayer, large numbers of capillaries and small glands produce visible; control group8-12Tiancheng fiber cell number than the experimental group was less, the presence ofa small amount of inflammatory cells, tissue structure is not clear.18days two groupsof the wounds healing, cellular species decreased, the existence of a large number ofcollagen bundles, capillary number, the basic recovery of epidermal structure; theexperimental group epidermal structure is relatively smooth. Transmission electronmicroscopic observation: experimental group15000times under the visible fibroblastvolume is large, well-formed, collagen secretion rich (Fig.3);40000times can be seenunder the cell content is rich, intracellular amounts of endoplasmic reticulum andmitochondria, ribosomes, the number of large volume, fibroblasts without edema,rupture (Figure4). The control group was15000times lower fibroblast collagensecretion, intracellular edema (Fig.5);40000times showed markedly reduced amountof rough endoplasmic reticulum, endoplasmic reticulum cell contents significantlyexpanded, less edema, vesicles, rough endoplasmic reticulum ribosomes amount little,mitochondrial number than the experimental group was less, part mitochondrial blurred(Figure6). PCNA immunohistochemical results: Third,5,8,12,18days, theexperimental group the positive expression of PCNA average higher than that of thecontrol group, third,5,8days after the positive expression rate was higher at three pointsin time. Statistical analysis confirmed fifth,8day of the experiment group and the control group the positive rate of P <0.05, third,12days when P <0.01, with statisticalsignificance. BCL-2immunohistochemical results: control group third,5,8,12day ofpositive expression was lower in experimental group. In the wound healing process ofBCL-2expression in3-8days most apparent, peaked at fifth days. The experimentalgroup fifth days when the positive expression of BCL-2, far higher than that of thecontrol group and the other at each time point. MMP-2Western blot A MMP-2detectvisible group the highest expression level; group B expression is somewhat low; groupC MMP-2is low; group D for local wound given by GM-CSF, MMP-2expression isminimal; group E MMP-2expression slightly less, and group C expression.Conclusion: Exogenous GM-CSF treatment, type2diabetic rat wound healingspeed, healing in good condition. GM-CSF treatment has multiple target effect,including: by acting on PCNA indirectly promote fibroblast proliferation, acceleratewound healing; activation of BCL-2inhibition of apoptosis, resistance to exogenousdamage; the inhibition of MMP-2on extracellular hyaline membrane decompositioneffect.