Hongsheng Liang

Allografts of the Acellular Sciatic Nerve and Brain-Derived Neurotrophic Factor Repair Spinal Cord Injury in Adult Rats

Objective We aimed to investigate whether an innovative growth factor-laden scaffold composed of acellular sciatic nerve (ASN) and brain-derived neurotrophic factor (BDNF) could promote axonal regeneration and functional recovery after spinal cord injury (SCI). Methods Following complete transection at the thoracic level (T9), we immediately transplanted the grafts between the stumps of the severed spinal cords. We evaluated the functional recovery of the hindlimbs of the operated rats using the BBB locomotor rating scale system every week. Eight weeks after surgery, axonal regeneration was examined using the fluorogold (FG) retrograde tracing method. Electrophysiological analysis was carried out to evaluate the improvement in the neuronal circuits. Immunohistochemistry was employed to identify local injuries and recovery. Results The results of the Basso-Beattie-Bresnahan (BBB) scale indicated that there was no significant difference between the individual groups. The FG retrograde tracing and electrophysiological analyses indicated that the transplantation of ASN-BDNF provided a permissive environment to support neuron regeneration. Conclusion The ASN-BDNF transplantation provided a promising therapeutic approach to promote axonal regeneration and recovery after SCI, and can be used as part of a combinatory treatment strategy for SCI management.

Reversal of P-glycoprotein-mediated multidrug resistance in vitro by milbemycin compounds in adriamycin-resistant human breast carcinoma (MCF-7/adr) cells

The effects of milbemycin A(4) (MB A(4)), milbemycin oxime A(4) (MBO A(4)) and milbemycin beta(1) (MB beta(1)) on reversing multidrug resistance (MDR) of tumor cells were firstly conducted according to the following research, including MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay, the accumulation of adriamycin, the accumulation and efflux of rhodamine 123 (Rh123), the regulations of MDR1 gene, and expression of P-gp. The three milbemycins (5muM) showed strong potency to increase adriamycin cytotoxicity toward adriamycin-resistant human breast carcinoma cells MCF-7/adr with reversal fold (RF) of 21.42, 19.06 and 14.89, respectively. In addition, the mechanisms of milbemycins on P-glycoprotein (P-gp)-mediated MDR demonstrated that the milbemycins significantly increased the intracellular accumulations of adriamycin and Rh123 via inhibiting P-gp transport function. Based on the analysis of the P-gp and MDR1 gene expression using flow cytometry and RT-PCR, the results revealed that milbemycin compounds, particularly MB A(4), could regulate down the expression of the P-gp and MDR1 gene. These findings suggest that the milbemycins probably represent promising agents for overcoming MDR in cancer therapy, and especially MB A(4) is better modulator with the lowest toxicity.

Transplantation of Bone Marrow Stromal Cells Enhances Nerve Regeneration of the Corticospinal Tract and Improves Recovery of Neurological Functions in a Collagenase-Induced Rat Model of Intracerebral Hemorrhage

The reorganization of brain structures after intracerebral hemorrhage (ICH) insult is crucial to functional outcome. Although the pattern of neuronal rewiring is well-documented after ischemic stroke, the study of brain plasticity after ICH has been focusing on the enhancement of dendritic complexity. Here we hypothesized that functional restoration after ICH involves brain reorganization which may be favorably modulated by stem cell transplantation. In this study, bone marrow stromal cells (BMSCs) were transplanted into the perilesional sites of collagenaseinduced ICH in adult rats one day after ICH injury. Forelimb functional recovery was monitored with modified limb placing and vibrissae-elicited forelimb placement tests. Anterograde and retrograde tracing were used to assess the reorganization of bilateral forelimb areas of the sensorimotor cortex. We found that in rats transplanted with BMSCs after ICH injury, axonal sprouting occurred in the contralateral caudal forelimb area of the cortex, and was significantly higher than in ICH rat models that received only the vehicle (P < 0.01). The number of positive neurons in the ipsilateral rostral forelimb area of the cortex of the BMSC group was 1.5-to 4.5-fold greater than in the vehicle group (P < 0.05). No difference was found between the BMSC and vehicle groups in hemispheric atrophy or labeled neurons in the ipsilateral caudal forelimb area (P = 0.193). Scores for improved functional behavior in the BMSC group were in accord with the results from histology. Neuronal plasticity of the denervated corticospinal tract at bilateral forelimb areas of the cortex in the collagenase-induced ICH rat models was significantly enhanced by BMSC transplantation. BMSC transplantation may facilitate functional recovery after ICH injury.

The effect of simvastatin treatment on proliferation and differentiation of neural stem cells after traumatic brain injury.

OBJECTIVE To study the effect of simvastatin on neurological functional recovery after traumatic brain injuries (TBI) and the possible molecular mechanisms, we evaluated simvastatin-induced proliferation and differentiation of neural stem cells (NSCs) in vitro and in vivo and possible involvement of Notch-1 signaling in this process. METHODS Adult Wistar rats were randomly divided into three groups (n=28 for each): sham group, saline-treated group and simvastatin-treated group. Simvastatin was given orally at a dose of 1mg/kg/day starting at day 1 after TBI. At 1, 3, 7, 14, 21, 28, and 35 days after simvastatin treatment, functional outcome was measured using modified neurological severity scores (mNSS). Immunofluorescence of nestin was used to identify neurogenesis of NSCs in injured area of TBI rats. Western blot was applied to detect the expression level of Notch-1 protein in TBI rats with simvastatin. RESULTS Immunostaining showed a significant increase in the number of nestin-positive cells in injured area of the simvastatin-treated group compared to that of the saline-treated group (p<0.05). In in vitro experiment, simvastatin induced enhanced proliferation and neurogenesis of cultured NSCs and elevated Notch-1 protein expression. Co-incubation of γ-secretase inhibitor, an inhibitor of Notch-1 pathway, with simvastatin abolished its neurorestoration effect. Most importantly, the simvastatin-treated group had significantly decreased mNSS at day 35 after TBI compared with the saline-treated group (p<0.05). CONCLUSION Simvastatin treatment enhanced neurological functional recovery after TBI possibly via activation of Notch signaling and increasing neurogenesis in the injured area.

Reversal of P-glycoprotein-mediated multidrug resistance in vitro by doramectin and nemadectin

Objectives Multidrug resistance (MDR) is a serious obstacle encountered in cancer treatment. This study was performed to explore the reversal of MDR by doramectin from the avermectin family and nemadectin belonging to the milbemycin family.

OPN gene polymorphism and the serum OPN levels confer the susceptibility and prognosis of ischemic stroke in Chinese patients.

Aim: To investigate the association of Osteopontin (OPN) gene polymorphism and serum thrombin-cleaved OPN level with the susceptibility to ischemic stroke (IS) and its prognosis. Methods: A total of 377 patients with IS and 551 healthy individuals were recruited. The OPN gene polymorphisms at -156 G>GG, -443 C>T and -66 T>G were genotyped. Serum full-length and the thrombin-cleaved OPN were determined. Results: We found that only the -443 C>T polymorphism was significantly associated with the susceptibility to IS. The -443 CC represented a near 2 time higher risk for IS incidence than TT carriers. Also, the -443 CC genotype had significantly poorer outcome and they significantly had higher occurrence for bad recovery as determined by modified Rankin Scale (mRS) (OR=2.18, p=0.043) and Barthel Index (BI) (OR=2.12, p=0.05). The mean serum thrombin-cleaved OPN level in IS group were significantly higher than that in control group. ROC analysis showed that the thrombin-cleaved OPN level (cut-off value, 166.8 ng/ml) can discriminate IS patients from controls with a specificity of 86.3% and a sensitivity of 57.7%. The serum thrombin-cleaved OPN was significantly associated with the clinical outcome at 12 months after discharge from hospital. Conclusion: These results suggest that the -443 C>T polymorphism of OPN gene and serum thrombin-cleaved OPN can be used as a biomarker for the susceptibility and prognosis of IS patients.

Reversal effects of two new milbemycin compounds on multidrug resistance in MCF-7/adr cells in vitro.

Development of agents to overcome multidrug resistance (MDR) is important in cancer chemotherapy, and the overexpression of P-glycoprotein (P-gp) is one of the major mechanisms of MDR. In this paper, we evaluated the effects of two new milbemycin compounds, milbemycin β(14) and secomilbemycin D, isolated from fermentation broth of S. bingchenggensis on reversing MDR of adriamycin-resistant human breast carcinoma (MCF-7/adr) cells. We observed that the both milbemycins (5 μM) showed strong potency to increase adriamycin cytotoxicity toward MCF-7/adr cells with reversal fold (RF) of 13.5 and 10.59, respectively. In addition, the mechanisms of milbemycins on reversing P-gp-mediated MDR demonstrated that they significantly increased the accumulations of adriamycin and Rh123 via inhibiting P-gp efflux in MCF-7/adr cells. Furthermore, the results also revealed that milbemycin β(14) and secomilbemycin D could regulate down the expression of P-gp, but not affect the expression of MDR1 gene. In conclusion, our observations suggest that the two new milbemycin compounds probably represent the promising agents for reversing MDR in cancer therapy.

TUSC3 suppresses glioblastoma development by inhibiting Akt signaling

Glioblastoma multiform is one of the most common and most aggressive brain tumors in humans. The molecular and cellular mechanisms responsible for the onset and progression of GBM are elusive and controversial. The function of tumor suppressor candidate 3 (TUSC3) has not been previously characterized in GBM. TUSC3 was originally identified as part of an enzyme complex involved in N-glycosylation of proteins, but was recently implicated as a potential tumor suppressor gene in a variety of cancer types. In this study, we demonstrated that the expression levels of TUSC3 were downregulated in both GBM tissues and cells, and also found that overexpression of TUSC3 inhibits GBM cell proliferation and invasion. In addition, the effects of increased levels of methylation on the TUSC3 promoter were responsible for decreased expression of TUSC3 in GBM. Finally, we determined that TUSC3 regulates proliferation and invasion of GBM cells by inhibiting the activity of the Akt signaling pathway.

Human amniotic epithelial stem cells inhibit microglia activation through downregulation of tumor necrosis factor-α, interleukin-1β and matrix metalloproteinase-12 in vitro and in a rat model of intracerebral hemorrhage

BACKGROUND AIMS The molecular mechanisms by which stem cell transplantation improves functional recovery after intracerebral hemorrhage (ICH) are not well understood. Accumulating evidence suggests that microglia cells are activated shortly after ICH and that this activation contributes to secondary ICH-induced brain injury. We studied the effect of human amniotic epithelial stem cells (HAESCs) on microglia activation. METHODS To study the effect of HAESCs in vitro, we used thrombin to activate the microglia cells. Twenty-four hours after thrombin treatment, the levels of tumor necrosis factor-α and interleukin-1β were measured by enzyme-linked immunosorbent assay. In vivo, the HAESCs were transplanted into the rat striatum 1 day after collagenase-induced ICH. The expression levels of matrix metalloproteinase (MMP)-12 and microglia infiltration in the peri-hematoma tissues were determined 7 days after ICH through the use of reverse transcriptase-polymerase chain reaction and immunohistochemical analysis, respectively. RESULTS Thrombin-activated microglia expression of tumor necrosis factor-α, interleukin-1β and MMP-12 was significantly reduced through contact-dependent and paracrine mechanisms when the HAESCs were co-cultured with microglia cells. After transplantation of HAESCs in rat brains, the expression levels of MMP-12 and microglia infiltration in the peri-hematoma tissues were significantly reduced. CONCLUSIONS Our observations suggest that microglia activation could be inhibited by HAESCs both in vitro and in vivo, which may be an important mechanism by which the transplantation of HAESCs reduces brain edema and ameliorates the neurologic deficits after ICH. Therefore, we hypothesize that methods for suppressing the activation of microglia and reducing the inflammatory response can be used for designing effective treatment strategies for ICH.

A novel animal model of partial optic nerve transection established using an optic nerve quantitative amputator.

Background Research into retinal ganglion cell (RGC) degeneration and neuroprotection after optic nerve injury has received considerable attention and the establishment of simple and effective animal models is of critical importance for future progress. Methodology/Principal Findings In the present study, the optic nerves of Wistar rats were semi-transected selectively with a novel optic nerve quantitative amputator. The variation in RGC density was observed with retro-labeled fluorogold at different time points after nerve injury. The densities of surviving RGCs in the experimental eyes at different time points were 1113.69±188.83 RGC/mm2 (the survival rate was 63.81% compared with the contralateral eye of the same animal) 1 week post surgery; 748.22±134.75 /mm2 (46.16% survival rate) 2 weeks post surgery; 505.03±118.67 /mm2 (30.52% survival rate) 4 weeks post surgery; 436.86±76.36 /mm2 (24.01% survival rate) 8 weeks post surgery; and 378.20±66.74 /mm2 (20.30% survival rate) 12 weeks post surgery. Simultaneously, we also measured the axonal distribution of optic nerve fibers; the latency and amplitude of pattern visual evoke potentials (P-VEP); and the variation in pupil diameter response to pupillary light reflex. All of these observations and profiles were consistent with post injury variation characteristics of the optic nerve. These results indicate that we effectively simulated the pathological process of primary and secondary injury after optic nerve injury. Conclusions/Significance The present quantitative transection optic nerve injury model has increased reproducibility, effectiveness and uniformity. This model is an ideal animal model to provide a foundation for researching new treatments for nerve repair after optic nerve and/or central nerve injury.