• Archives of Reconstructive Microsurgery
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• v.6 no.1
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• pp.9-28
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• 1997

Adequate vascularization is pivotally essential for a successful nerve graft. Theoretically, the immediate vascularization will inhibit fibroblast infiltration and stimulate nerve cell regeneration. In this study, histomorphological and electrophysiological studies were performed to determine if vascularized grafts are functionally superior. In rat model, a 4cm segment of the sciatic nerve was obtained and placed as a non vascularized graft on one side, and as a vascularized graft connected to the inferior gluteal vessels on the opposite side. To determine the compound action potential of the gastrocnemius muscle, electromyography was done after 2, 3 and 4 months. Histomorphologically, the distribution of myelinated nerve fibers and Schwann cell were evaluated after toluidine blue staining, The following resutls were obtained: 1. The electrophysiological studies showed no difference between the nonvascularized and vascularized grafts. 2. Two and three months after grafting, myelinated nerve fibers were more abundant in the vascularized proximal, middle and distal areas in all nerve fibers of varying diameters. 3. In the post-nonvascularized graft 2-month group, a few myelinated nerve fibers were present in the proximal and middle areas, but none distally. In the post-vascularized graft 2 month group, myelinated nerve fibers ranging $2-8{\mu}m$ were present in all three areas. 4. In the post-nonvascularized graft 3 month group, a few myelinated nerve fibers ranging in $2-6{\mu}m$ were present in all three areas, but in the post-vascularized graft 3 month group, many myelinated nerve fibers ranging in $2-10{\mu}m$ were present in all three areas. 5. In the post-graft 4-month group, more myelinated nerve fibers were present in all three areas of the vascularized grafts. However, nerve fibers of less than $2{\mu}m$ in diameter were more abundant in the non vascularized grafts. 6. Schwann cells were more abundant in the proximal, middle and distal areas of the post-vascularized 2, 3 and 4-month grafts. Based on these findings, the immediate restoration of circulation in vascularized nerve grafts allows for the increased number of surviving Schwann cells, rapid healing of the axon and myelin sheath changes which occur during Wallerian degeneration, and thus is able to stimulate a morphologically optimal regeneration.

• Archives of Plastic Surgery
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• v.33 no.2
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• pp.205-212
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• 2006

Using adipose derived stem cells(ASCs), neurogenic differentiation was induced in a mono layered culture medium containing neuronal induction agents. Cells differentiated to the neuronal cells were observed with a inverted microscope and immunofluorecent study. We made a 15 mm long defect in the sciatic nerve of 14 rats and connected a silicone tube to the defect. Then, we mixed neuronal progenitor cells differentiated from ASCs with collagen gel and grafted them to a group of rats(experimental group) and grafted only collagen gel into another group(control group). In 4 and 8 weeks after the graft, histological observation was made. According to the result, the number and diameter of myelinated axons were significantly increased in the experimental group. In addition, the nerve conduction velocity was improved more in the experimental group and neovascularity also increased. Moreover, reaction with S100 and p75 was observed in regenerated nerves in the experimental group, suggesting that the grafted cells were differentiated into supportive cells such as Schwann's cells. In conclusion, this research proved that ASCs can multiply and differentiate into neuronal cells. If they are grafted into nerve defects, the grafted cells are differ entiated into supportive cells such as Schwann's cells and thus contribute to nerve regeneration. Accordingly, the use of adipose tissue obtained easily without the limitation of donor site can be greatly helpful in treating peripheral nerve defects.

• Polymer(Korea)
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• v.30 no.5
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• pp.445-452
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• 2006

In this study, we investigated interaction of Schwann cells (SCs) with various cell-adhesive coated polymer surface. We used cell-adhesives that like a fibronectin (FN), fibrinogen(FG), laminin(LM), vitronectin (VN), poly-D-Iysine (PDL), and poly-L-Iysine (PLL) to coat PLGA film surface and evaluated the surface property of coated or not PLGA films by measurement of water contact angle and ESCA. SCs were cultured on coated or non-coated PLGA film surface, and then examined the cell adhesion and proliferation by cell count and SEM observation. Cell count results revealed initial cell adhesion related to protein adsorption on PLGA surface. In addition, serum content in media related to cell proliferation rate. In this result, we recognized that adhesion and proliferation of SCs were affected by specific cell-adhesives. In these results, we recognized that is important to provide the suitable surface environment according to cell types and culture condition for improvement of cell adhesion and proliferation.

• Polymer(Korea)
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• v.33 no.2
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• pp.118-123
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• 2009

Keratin contains regulatory molecules that can enhance neuronal cell activity. We fabricated keratin/ PLGA films using 0, 10, 20, and 50 wt% of keratin using solvent casting method. We measured the contact angle of each film and cell proliferation was assayed by counting the cells attached on the film. Adhered cell morphology was confirmed by scanning electron microscope. RT-PCR was conducted to evaluate the gene expression of NF, NSE, and S-100, the Schwann cell markers. The keratin content of 20 and 50 wt% provided higher wettability than PLGA. The 20 wt% keratin was better in cell adhesion and proliferation of SCs than other keratin/PLGA films. The phenotypic stability of SC was maintained with the keratin content of 10 and 20 wt%.

• The Korean Journal of Pharmacology
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• v.26 no.2
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• pp.177-183
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• 1990

This study was undertaken to confirm whether or not the sympathetic nervous system takes part in the liver regeneration after partial hepatectomy. The male Sprague-Dawley rats were pretreated with I.P. injection of guanethidine 25 mg/kg: single dose (G-1); multiple doses once a day for 3 days (G-3), for 5 days (G-5), or for 6 days (G-6). The rats were subjected to partial hepatectomy $(70.4{\pm}1.99%)$ under light anesthesia of diethyl ether. 1) The systolic blood pressure of control rat was $98.0{\pm}3.9\;mmHg$ and was not affected by G-1. But after the pretreatment with G-3, G-5 or G-6, the pressure was markedly decreased by over 25 %. 2) Both of plasma norepinephrine and epinephrine levels showed the marked increases 3 hrs after the hepatectomy. However, the increases are entirely inhibited by G-1 or G-6. 3) All the liver contents of putrescine, spermidine and spermine showed the significant increases 6 hrs after the hepatectomy and were not affected by G-1 or G-6 with the exception of the inhibition of putrescine increase by only G-6. The present results suggest that the sympathetic activation appeared after partial hepatectomy seems not to play an important role in rat liver regeneration.

• The Journal of Korean Physical Therapy
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• v.17 no.4
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• pp.589-600
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• 2005

The purposes of this study is to discuss and analyze the effect on the recovery from cut in sciatic nerve. This study used 9 weeks male rats of Sprague-Dawley family. Rat in groups 4 were treated with pulsed therapeutic ultrasound for 3 minutes. 3 times weekly at 3MHz respectively (intensity; $0.2W/cm^2,\;0.5W/Cm^2,\;10W/cm^2$); rat in group 1 received placebo ultrasound. In addition, changes of serum aspartate amino-transferase(AST) and creatine phosphokinase(CPK) levels were also demonstrated with diameter of individual muscle fasciculate and number of muscle fiber in each of three types of muscles located in gastrocnemius, soles. The results of comparing the changes in groups are as follows; 1. We found out that hypertrophic epineurium was present in sciatic nerve injured ultrasound treatment of groups. 2. In the gastrocnemius morphological investigation of the group I (control group), severe muscle atrophy were observed at the 7th days of the sciatic nerve injury. however, muscle atrophy of the group IV ($1.0W/cm^2$) were slightly recovered at the 14th days after treatment ultrasound. At the 28th days, muscular fibers were formed in polygon and were significantly recovered. 3. C-fos immunoreactive of the group II ($0.2W/cm^2$), III ($0.5W/cm^2$) were remarkably increased at the 1th day after treatment of ultrasound. Group IV were markedly deceased. 4. Brain-Derived Neurotrophic Factor(BDNF) immunoreactive of the group II, III were increased after 7 days of the sciatic nerve injury. Group IV were markedly increased from 14th days to 28th days after treatment of ultrasound. 5. A significant increase of serum AST levels were demonstrated in control group. However, serum AST levels of massage groups were significantly decreased compared to that of control group in followed order ; ($0.2W/cm^2<0.5W/cm^2<1.0W/cm^2$). 6. A significant increase of serum CK levels were demonstrated in control of group. However, serum CK levels of massage groups were significantly decreased compared to that of control group in followed order ; ($0.2W/cm^2<0.5W/cm^2<1.0W/cm^2$). The above results suggest that ultrasound treatment after peripheral nerve injury might reduce noxious stimuli, facilitate nerve recovery and effective in the functional improvement delaying muscle atrophy or degeneration.

• Journal of Physiology & Pathology in Korean Medicine
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• v.26 no.4
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• pp.462-468
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• 2012

The present study was conducted with the object of examining how ST36 and LR3 electro acupuncture affects the injured side and the intact side in rats with sciatic nerve injury. For this study, we divided rats into a control group that were injured but not treated, experimental group I that were injured and had electro acupuncture on the intact side, experimental group II that were injured and had electro-acupuncture on the injured side, and experimental group III that were injured and had electro-acupuncture on both the intact side and the injured side, and performed behavioral and immunohistochemical tests. The results of this study are as follows. In the results of hot plate test, on Day 1 of experiment, experimental group I did not show a statistically significant difference but experimental group II and III showed a statistically significant difference, and on Day 2 and 3, all of experimental group I, II and III showed a statistically significant difference. In the results of SFI test, on Day 7 of experiment, experimental group I did not show a statistically significant difference but experimental group II and III showed a statistically significant difference, and on Day 14 and 21, all of experimental group I, II and III showed a statistically significant difference. In the results of immunohistochemical test, the expression of c-fos decreased gradually on Day 1, 2 and 3 in all the experimental groups, and the decrease was larger in the experimental groups that had electro-acupuncture than in the control group. In the results of immunohistochemical test, the expression of BDNF increased gradually on Day 7, 14 and 21 in all the experimental groups, and the expression was higher in the experimental groups than in the control group. Summing up the results, ST36 and LR3 electro acupuncture was effective when it was applied to the injured side and to both sides after sciatic nerve injury, but it was also effective in pain relief and nerve regeneration when it was applied to the intact side.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.27 no.5
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• pp.446-456
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• 2005

Nerve growth factor(NGF) has a critical role in peripheral nerve regeneration. The aim of this study is to construct a well-functioning hNGF-$\beta$ recombinat adenovirus for the ultimate development of improved method to promote peripheral nerve regeneration with adenovirus mediated hNGF-$\beta$ gene transfection into Schwann cells. First PCR associated cloning of GFP-tagged hNGF-$\beta$ which was ligated into E1/E3 deleted adenoviral vector was performed and tranfected into E. coli to construct hNGF-$\beta$ recombinant adenovirus. After production of recombinat adenovirus in a large scale, its transfection efficiency, expression, and function were evaluated using cell lines or primarily cultured cells of HEK293 cells, Schwann cells, fibroblast(NIH3T3) and myocyte(CRH cells). GFP expression was observed in 90% of infected cells compared to uninfected cells. Total mRNA isolated from hNGF-$\beta$ recombinat adenoviru infected cells showed strong RT-PCR band, however, LacZ recombinant adenovirus infected or uninfected cells did not. NGF quantification by ELISA showed a maximal release of 18.865 +/- 0.31ng/mL at 4th day. PC-12 cells exposed to media with hNGF-$\beta$ recombinant adenovirus infected Schwann cell demonstrated higher levels of differentiation compared with controls. We generated hNGF-$\beta$ recombinant adenovirus and induced over expression of NGF successfully in nonneuronal and neuronal cells. Following these result, it is expected to develop an improved treatment strategy peripheral nerve regeneration using the hNGF-$\beta$ gene transfected cells.

• Archives of Plastic Surgery
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• v.34 no.3
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• pp.279-284
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• 2007

Purpose: The vein graft was considered as a useful conduit for nerve defect. But the problem is that it might be collapsed in long vein graft state. A new experimental model using vein graft filled with hyaluronic acid was considered. Methods: Thirty rats were used for the experimental animal. In group I, one side of the femoral nerve was exposed and a segment was removed about 15mm. The neural gap was connected with nerve graft. In group II, the nerve gap was connected with vein graft only. In group III, the nerve gap was connected with vein graft filled with hyaluronic acid. A walking track analysis was made periodically for 2 months and NCV(nerve conduction velocity) was executed at the end of the experiment. And morphologic studies were also done for all groups Results: In a walking track analysis, the toe-spread was widen and the foot-length was lengthened. The recovery of the toe-spread and foot length was checked 2 weeks interval, periodically for two months. The SFI (sciatic function index) was $-52.5{\pm}8.2$ in group I, $-68.1{\pm}4$ in group II, $-55.3{\pm}7.9$ in group III. In electrophysiological study, NCV(nerve conduction velocity) was $26.71{\pm}3.11m/s$ in group I, $17.94{\pm}4.35m/s$ in group II, $25.69{\pm}2.81m/s$ in group III. The functional recovery in group I and III was superior to that the group II statistically(p < 0.05) Under electromicroscopic study, the number of the myelinated axons were $1419.1{\pm}240$ in group I, $921.7{\pm}176.8$ in group II, $1322.2{\pm}318$ in group III. The number of the myelinated axons were much more in group I and III than group II statistically (p<0.05). Conclusion: This study suggested that the vein graft filled with hyaluronic acid is more effective than vein graft only for the conduit of the nerve gap. It was thought that the technique could be used in clinical cases with nerve defects as an alternative method to classical nerve grafts.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.29 no.3
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• pp.151-156
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• 2003

This study was performed to develop a useful nerve conduit which provides favorable environment for Schwann cell viability and proliferation. Milipore membrane of $0.45{\mu}m$ pore size was selected because it permits nutritional inflow from the outside of the conduit and prevents from invading the fibrotic tissue into the conduit. The membrane was rolled and sealed to form a conduit of 2mm diameter and 20mm length. To improve the axonal regeneration and to render better environment for endogenous and exogenous Schwann cell behaviour, the microgeometry and surface of conduit was modified by coating with thin film of calcium phosphate. Cellular viability within the conduit and attachment to its wall were assessed with MTT assay and SEM study. Milipore filter conduit showed significantly higher rate of Schwann cell attachment and viability than the culture dish. However, the reverse was true in case of fibroblast. Coating with thin film of low crystalline calcium phosphate made more favorable environment for both cells with minimal change of pore size. These findings means the porous calcium phosphate coated milipore nerve conduit can provide much favorable environment for endogenous Schwann cell proliferation and exogenous ones, which are filled within the conduit for the more advanced strategy of peripheral nerve regeneration, with potential of reducing fibrotic tissue production.