Immunohistochemical profile of transforming growth factor-ß1 and basic fibroblast growth factor in sciatic nerve anastomosis following pinealectomy and exogenous melatonin administration in rats

PubMed ID: 16831553Collagen scar formation at the cut end of a peripheral nerve, an important problem in clinical practice for neurosurgeons, obstructs sprouting of axons into appropriate distal fascicles, and thereby limits the regeneration process. Researchers have attempted to control collagen accumulation and neuroma formation with various physical and chemical methods, but with limited functional success. Recently, it has been demonstrated that transforming growth factor (TGF)-ß and basic fibroblast growth factor (bFGF) play an important role in collagen production by fibroblasts and in Schwann cell activity. In our study, rats were divided into a control group, a melatonin-treated group, a surgical pinealectomy group, and a group treated with melatonin following pinealectomy. They then underwent a surgical sciatic nerve transection and primary suture anastomosis. At 2 months after anastomosis, the animals were sacrificed and unilateral sciatic nerve specimens, including the anastomotic region, were removed and processed for immunohistochemical study from two animals in each group. For each antibody, immunoreactivity was assessed using a semiquantitative scoring system. Strong TGF-ß1 and/or bFGF expression was observed in the epineurium of animals that underwent pinealectomy, but no or weak staining was observed in animals in the control and melatonin treatment groups. Based on these data, we suggest that both TGF-ß1 and bFGF have important roles in control of collagen accumulation and neuroma formation at the anastomotic site, and that the pineal neurohormone melatonin has a beneficial effect on nerve regeneration. © 2006 Elsevier Ltd. All rights reserved.The authors are grateful to Professor S. Kaplan for helpful discussions and M. Koltas for linguistic help. We also extend our thanks to Mr. B. Ata, Mr. Ç. Uz, and Mrs. S. Balkız for their skilful technical assistance. This work was supported by a grant from the Scientific and Technical Research Council of Turkey (TUBITAK), Grant No. SBAG-2323 (100S075). This study was presented in part at the 4 th Asian-Pacific International Congress of Anatomists (APICA), 7–10 September 2005, Kusadasi, Turkey. -

Assessment of effects of pinealectomy and exogenous melatonin administration on rat sciatic nerve suture repair: An electrophysiological, electron microscopic, and immunohistochemical study

PubMed ID: 15565477Background. Collagen scar formation at the cut end of a nerve, an important problem in clinical practice for neurosurgeons in peripheral nerve surgery, obstructs sprouting of axons into appropriate distal fascicles, and thereby limits nerve regeneration. Researchers attempt to control collagen accumulation in the formation of neuroma by various physical and chemical methods, but these have yielded only limited functional success. This is the first experimental study investigating the effects of melatonin (MLT) on nerve repair and neuronal regeneration in rat sciatic nerve suture repair. Methods. The hypothesis that exogenous MLT administration may inhibit the formation of neuroma in peripheral nerve surgery was investigated in rat sciatic nerve model. In this study, a total of 80 rats were used for control groups (Groups Ia, Ib, IIa, and IId), MLT group (Group Ic), surgical pinealectomy (Px) groups (Groups IIb and IIc), and group of MLT treatment following Px procedure (Group IIe). All animals underwent a surgical intervention consisting of bilateral sciatic nerve section and primary suture repair. At 8 weeks after repair, the animals were killed following completion of recording of nerve action potentials (NAPs). Then, unilateral sciatic nerve specimens including the suture repair region were carefully removed and the excised segments were processed for electron microscopy examination. Afterwards, contralateral sciatic nerve specimens from two animals from each group were removed and stained for immunohistochemical analysis. Results. Results of morphometric analysis revealed that Px procedure caused an elevation of collagen content of the sciatic nerve and macroscopic neuroma formation, and that there was a statistically significant reduction in collagen content of the same region in pinealectomized animals treated with MLT (p < 0.001). Accordingly, electrophysiological findings demonstrated that the stimulus intensities required to excite a NAP response were increased in surgical Px group, but the presence of a reduced threshold response was found in the group treated with MLT following Px procedure (p < 0.01). Immunohistochemical staining for Type I collagen and Type III collagen was markedly more intense in the epineurium of animals after Px. Virtually no or only weak staining was observed in animals in control groups and the MLT treatment group. Results of immunohistochemical analysis revealed that surgical Px procedure caused a strong immunoreactivity for Type I collagen and Type III collagen in all connective tissue planes of the nerve, especially in the epineurium, and there was a statistically significant reduction in immunoreactivity of the repair region in animals receiving MLT treatment after Px procedure (p < 0.001). Conclusion. This study demonstrates that exogenous MLT administration significantly inhibits collagen accumulation in the formation of neuroma in the suture repair site and thereby improves nerve regeneration. From a clinical standpoint, the positive effect of MLT administration on neuroma formation and nerve regeneration seems a particularly attractive treatment option. Therefore, we believe that nerve repair with addition of MLT may be a worthwhile option in addition to other treatment modalities in case of MLT deficiency, such as aging. However, further experimental and clinical studies using functional analysis warranted to confirm this result in future. © Springer-Verlag 2004

Evaluation of the relationship between inducible nitric oxide synthase (iNOS) activity and effects of melatonin in experimental osteoporosis in the rat

WOS: 000237091700008PubMed ID: 16362227Inducible nitric oxide synthase (iNOS) plays a critical role in the pathogenesis of osteoporosis. iNOS generates nitric oxide (NO), a free radical contributing to the imbalance between bone formation and resorption caused by estrogen depletion. Melatonin is the major product of the pineal gland which is known to diminish iNOS expression and NO production significantly. The aim of this study was to determine the distribution of iNOS and the amount of apoptotic cells after melatonin treatment in ovariectomized rats. Since previous studies have shown that constitution of bone formation is primarily sustained in nucleus pulposus and epiphyseal cartilage, experiments were carried out on nucleus pulposus and epiphyseal cartilage; additional quantitation of osteoblasts and osteoclasts were evaluated on vertebral area as well. Vertebral sections of ovariectomized rats were obtained from formalin-fixed and parafin-embedded blocks. iNOS expression and quantitation of apoptotic cells in nucleus pulposus and epiphyseal cartilage were evaluated using indirect immunoperoxidase and TUNEL techniques, respectively. The number of osteoclasts and osteoblasts in trabecular bone was determined using histomorphometry. Ovariectomy increased iNOS expression and the number of apoptotic cells in nucleus pulposus and epiphyseal cartilage, whereas a 4-week treatment with melatonin (10 mg/kg/day) resulted in the reduction of both effects. These data indicate that there is strong influence of melatonin application on expression of iNOS, apoptosis, osteoclast and osteoblast numbers after ovariectomy. In conclusion, melatonin besides its usual use as an antiaging hormone, may also be an effective hormone in treatment of bone changes in estrogen deficiency states