Gulgun Oktay

Matrix metalloproteinase‐2 and ‐9 activities in human keloids, hypertrophic and atrophic scars: a pilot study

Proteolytic degradation of extracellular matrix is one of the principal features of cutaneous wound healing but little is known about the activities of gelatinases; matrix metalloproteinase‐2 (MMP‐2) and matrix metalloproteinase‐9 (MMP‐9) on abnormal scar formation. The aim of this study is to determine collagen levels and the gelatinase activities in tissue from hypertrophic scars, atrophic scars, keloids and donor skin in 36 patients and 14 donors. Gelatinase levels (proenzyme + active enzyme) were determined by ELISA and their activities by gelatin zymography. MMP‐9 activity was undetectable in gelatin zymography analysis. Pro‐MMP‐2 levels (median) were highest in normal skin group 53.58 (36.40–75.11) OD µg−1 protein, while active MMP‐2 levels were highest in keloid group 52.53 (42.47–61.51) OD µg−1 protein. The active/pro ratio was the highest in keloid group 0.97 followed by hypertrophic scar, normal skin and atrophic scar groups 0.69 > 0.54 > 0.48, respectively. According to results of our study, the two‐phase theory of the duration of hypertrophic scar and keloid formation can be supported by the data of tissue collagen and gelatinase analysis. This study is the first to relate scar formation relationship in regard to gelatinase activation ratio in a keloid, hypertrophic and atrophic scar patient group which is chosen appropriate in age and sex. Copyright

EFFECTS OF GRANULOCYTE-MACROPHAGE COLONY -STIMULATING FACTOR ON INCISIONAL WOUND HEALING IN AN EXPERIMENTAL DIABETIC RAT MODEL

The exact nature of poor wound healing in diabetes is uncertain. Neutrophils play a critical role in the host defense mechanism, and it is suggested that impaired neutrophil functions cause healing difficulties with or without infections in diabetic patients. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is used clinically when given systematically to increase the circulating neutrophils, but its wound-healing effects have not been systematically studied. This study was undertaken to examine the effects of GM-CSF on incisional wound healing in an experimental diabetic rat model. Forty rats were randomly divided into three groups, group I receiving saline as control, diabetes-induced group II receiving saline and diabetes-induced group III receiving GM-CSF. The anesthetized rats in all groups were wounded 21 days after diabetes induction by streptozotocin. Blood neutrophil counts and neutrophil fractions were also determined three days after wounding. Tensile strengths of wounded skin and the hydroxyproline (hyp) level of the wound were determined and wound healing processes were evaluated by light and electron microscopy, fourteen days after wounding. Neutrophil counts and phagocytosis were significantly increased in group III and neutrophil counts decreased in group II (p < 0.05). Although the hydroxyproline level of wound tissue significantly decreased in group II as compared with group III (p < 0.05), there was no differences of tensile strength between group II and III (p < 0.05). Wound score in group II was less than that in groups I and III (p < 0.05). It is concluded that PMN may have a role in modulating wound healing. GM-CSF may be useful for creating better wound healing healing. GM-CSF may be useful for creating better wound healing in risky patients such as diabetics.

The Relationship between Neutrophils and Incisional Wound Healing

The systemic administration of granulocyte-macrophage colony-stimulating factor (GM-CSF) is used clinically to increase circulating neutrophils, but its wound healing effects after intraperitoneal treatment have not been studied yet. We planned to investigate the effect of neutrophils on wound healing under cyclophosphamide and GM-CSF treatment. Forty rats were divided into three groups: control group (group I, n = 12) receiving saline, group II (n = 14) receiving cyclophosphamide and group III (n = 14) receiving GM- CSF. The rats in all groups underwent incisional wounding and were euthanized after 7 days. Blood neutrophil counts and functions, tensile strengths and the hydroxyproline level of skin were determined, and a histopathological evaluation of healing was made. Neutrophil counts and phagocytosis significantly increased in group III and decreased in group II. Although the skin hydroxyproline level did not differ, there was a difference in tensile strength of the wounded skin between group II and group III. The wound score in group II was lower than that in groups III and I. As a result we suggest that systemically given GM-CSF – by increasing the neutrophil count and neutrophil phagocytosis index – can enhance the tensile strength of surgical incisions.

Intrathecal sICAM-1 production in multiple sclerosis Correlation with triple dose Gd-DTPA MRI enhancement and IgG index

In this study the aim was to evaluate the intrathecal sICAM-1 production in multiple sclerosis (MS) patients during relapse and remission. In addition to this, we assessed whether there is a correlation between intrathecal sICAM-1 production and other disease activity markers such as IgG index and gadolinium enhancement in magnetic resonance imaging (MRI). Twenty four relapsing- remitting MS patients were included in the study. Serum and cerebrospinal fluid (CSF) samples were obtained both during relapse and remission. The soluble form of ICAM (sICAM) was measured by the ELISA method in serum and CSF. Cranial MRI with triple dose gadolinium injection was performed for each patient both during relapse and remission. Serum levels of sICAM-1 (245.23±92.88 ng/ml) were higher during relapse than those in remission (219.90±110.94 ng/ml), but the difference was not statistically significant. In relapse periods CSF levels of sICAM-1 (1.304±0.92 ng/ml) were higher than those in remission (1.06±0.86 ng/ml), but this was not significant. However, during relapse periods patients had significantly higher sICAM-1 index values (1.76±0.60) than those found during remission periods (1.01±0.44) (p<0.05). The IgG index values were higher in relapse periods than in remission (0.88±0.37 vs. 0.67±0.28) (p<0.005). On T1 weighted images following triple dose Gd injection, at least two or more enhancing lesions were present in 22/24 of the patients (91 %) in relapse and 4/24 of the patients (19 %) in remission. There was strong correlation both between the sICAM-1 index and Gd enhancement (r=0.72 p<0.05) and sICAM-1 index and IgG index in relapse (r=0.69 p<0.05). In conclusion, there is association between high sICAM-1 and IgG indices, as well as between high sICAM-1 index and Gd enhancing MRI lesions in relapsing MS patients.

Improvement of Colonic Healing by Preoperative Rectal Irrigation With Short-Chain Fatty Acids in Rats Given Radiotherapy

BACKGROUNDWe investigated the effect of preoperative rectal irrigation with short-chain fatty acids on irradiated colonic anastomosis in rats.METHODSSixty male Wistar rats were divided into four groups. Group I (control group, n = 15) underwent left colon resection and primary anastomosis. Group II (Short-chain fatty acids pretreatment group, n = 15) had short-chain fatty acids rectal irrigation for five days preoperatively. Group III (preoperative radiotherapy group, n = 15) underwent irradiation to the whole pelvis eight and four days before the operation, for a total dose of 20 Gy. Group IV (preoperative radiotherapy group + short-chain fatty acids pretreatment group, n = 15) had rectal irrigation with short-chain fatty acids for five days after the second irradiation. Within each group, animals were anesthetized to assess the clinical, mechanical, histologic, and biochemical parameters of anastomotic healing on either the third or seventh postoperative days.RESULTSThe mean bursting pressure was significantly low in Group III on Day 3 and was significantly high in Group IV on Day 7 (P = 0.001, P = 0.021). The burst occurred at the anastomoses in all animals tested on the third postoperative day, and outside of the anastomoses in all animals tested on the seventh postoperative day. The histologic parameters of anastomotic healing, such as epithelial regeneration and formation of granulation tissue, were significantly improved by use of preoperative rectal irrigation with short-chain fatty acids on Day 7. The amount of total and salt-soluble collagen concentrations significantly increased in Group IV compared with the control group on Day 3 (P = 0.008, P = 0.004).CONCLUSIONSome mechanical and histologic aspects of colonic anastomotic healing can be adversely affected by preoperative radiotherapy, but rectal irrigation with short-chain fatty acids may improve anastomotic healing.

Protective Effects of Resveratrol against Chronic Immobilization Stress on Testis

Objective. The aim of this study was to investigate protective effects of resveratrol, a strong antioxidant, against possible negative effects of chronic immobilization stress on testes of male rats histochemically, immunohistochemically, ultrastructurally, and biochemically. Material and Methods. Male Wistar rats were divided into 4 groups (n = 7). Group I, control group (C), was not exposed to stress. Group II, stress group (S), was exposed to chronic immobilization stress. In Group III, low dose resveratrol + stress group (LRS), rats were given 10 mg/kg/day resveratrol just before the stress application. In Group IV, high dose resveratrol + stress group (HRS), rats were given 20 mg/kg/day resveratrol just before the stress application. For chronic immobilization stress application animals were put in the plastic tubes (6 cm in diameter, 15 cm in length) during 32 days for 6 hours. All animals were sacrificed 18 hours after the last stress application. Results. Histochemical and ultrastructural investigations showed that in stress group there was germ cell deprivation in seminiferous tubules and increase of connective tissue on interstitial area. No significant changes were seen in low and high dose resveratrol groups. After immunohistochemical investigations, TUNEL (+) and Active Caspase-3 (+) cells were increased in seminiferous tubules of stress group compared with those control group, but they were decreased in low and high dose resveratrol groups. According to biochemically results, MDA, GSH, and testosterone levels in stress group showed no significant difference when compared with those of the other groups. Conclusion. The chronic immobilization stress increases oxidative stress and apoptosis and causes histological tissue damages; resveratrol can minimize the histological damage in testes significantly.

Pioglitazone Reduces Peritoneal Fibrosis via Inhibition of TGF-β, MMP-2, and MMP-9 in a Model of Encapsulating Peritoneal Sclerosis

Abstract Objective: Matrix metalloproteinases (MMPs) and transforming growth factor beta (TGF-β) were increased in peritoneal dialysis patients with encapsulating peritoneal sclerosis (EPS) and in chlorhexidine gluconate (CG)-induced peritoneal sclerosing animal models. Peroxisome proliferator-activated receptors (PPARs) are the major regulators of key metabolic pathways of various inflammatory responses in fibrosing processes in most tissues. The objective of this study was to investigate the effect of pioglitazone (Pio), a synthetic PPAR-γ ligand, on the development of peritoneal fibrosis in CG-induced EPS rats. Methods: Thirty-two Wistar albino rats were intraperitoneally injected with saline (C group n = 8) or with CG (1.5 mL/100 g; CG group, n = 8). Pio (30 mg/kg/day) was administered orally to another group of CG injected rats (the CG + Pio group, n = 8) and to another control group (Pio group, n = 8) from initiation to the end of this study. After 14 days of Pio administration, the rats were killed and the parietal and visceral peritoneum were harvested. TGF-β, MMP-2, MMP-9, tissue inhibitor of metalloproteinases (TIMP)-1, and TIMP-2 activity assays and a morphological examination of the peritoneal tissues were performed. Results: Pio significantly inhibited thickening of the submesothelial layer, fibrosis, and inflammation in the peritoneum. It also prevented increases in pro-MMP-2, pro-MMP-9, TIMP-1, and TGF-β activities. Conclusion: Pio, via MMP and TGF-β inhibition, may lessen accumulation of peritoneal extracellular matrix and fibrosis to some extent in an EPS model and might be a new approach to the amelioration of EPS.

Taurine inhibits increased MMP-2 expression in a model of oxidative stress induced by glutathione depletion in rabbit heart.

Matrix metalloproteinase enzymes (MMPs) activated by oxidative stress are involved in the pathogenesis of cardiovascular diseases. Glutathione (GSH) plays an important protective role against oxidatively induced damage in mammalian tissues. We investigated the possible role of gelatinases and the effect of the semiessential amino acid 2-aminoethanesulfonic acid (taurine) in oxidatively induced damage by GSH depletion in rabbit cardiac tissues. Rabbits were treated with buthionine sulfoximine (BSO), an effective GSH-depleting compound. BSO treatment significantly reduced GSH and increased MDA (malondialdehyde) levels. BSO treatment caused significant increase in proMMP-2 levels. MMP-9 (pro and active) expressions were not found in either treated- or untreated heart tissues. TIMP-1(endogenous inhibitor of MMP-9) and MT-MMP1 (endogenous activator of MMP-2) were not affected by BSO. Immunoscoring showed that MMP-2 expression significantly increased in hearts from BSO treated group but MMP-9 antibody did not show any significant positive immunostaining from all groups. Type I procollagen and total collagen did not significantly alter in heart tissues from all treatment groups. Taurine restored the increased MDA and the diminished GSH levels by BSO treatment. Pro MMP-2 expression was prevented by taurine. These results suggest that MMP-2 is a major gelanitase in rabbit hearts under oxidative stress and pharmacological inhibition of MMP-2 activation by taurine could represent a useful strategy for the prevention and/or treatment of different cardiovascular disorders.

Role of gelatinases (matrix metalloproteinases 2 and 9), vascular endothelial growth factor and endostatin on clinicopathological behaviour of rectal cancer.

Aim  The aim of this study was to evaluate the role of matrix metalloproteinases (MMPs), their tissue inhibitors [tissue inhibitors of metalloproteinases (TIMPs)] and activators [membrane‐type MMPs (MT1‐MMPs)], vascular endothelial growth factor (VEGF) and endostatin on clinicopathological variables and prognosis in patients with rectal cancer.

The role of endothelin receptor antagonism in collar‐induced intimal thickening and vascular reactivity changes in rabbits

Intimal thickening, due to smooth muscle cell migration and proliferation, is considered to be one of the major components of vascular proliferative disorders such as atherosclerosis and restenosis. One experimental model, resulting in intimal thickening in the rabbit, involves placing a silicon collar around the carotid artery, and is used in this study. Endothelin is known to act as a strong mitogen and to stimulate smooth muscle cell proliferation and migration. We investigated the contribution of endothelin to the development of collar‐induced intimal thickening and the effects of TAK‐044, (5mg kg−1 daily, s.c.), a non‐selective ETA/ETB receptor antagonist, on intimal thickening and vascular reactivity changes in the collared rabbit carotid artery. Endothelin levels and the intimal cross‐sectional area, as well as the ratio of intimal area to media (index), increased significantly in collared arteries as compared with those in sham‐operated arteries. TAK‐044 significantly inhibited intimal thickening and also decreased the index without affecting increased endothelin levels in collared arteries. Vascular reactivity changes in response to collaring produced predictable effects, such as decreased contractile responses to vasoconstrictor agents and increased sensitivity to serotonin (5‐hydroxytrypt‐amine, 5‐HT). In terms of contractile responses in this model, TAK‐044, in particular, did not affect collar‐induced vascular reactivity changes. These results suggest that endothelin may be involved in the pathogenesis of collar‐induced intimal thickening. As an endothelin receptor antagonist, TAK‐044 may potentially be beneficial in the treatment of atherosclerosis.