The protective role of tacrine and donepezil in the retina of acetylcholinesterase knockout mice

AIM:To determine the effect of different concentrations of the acetylcholinesterase (AChE) inhibitors tacrine and donepezil on retinal protection in AChE^+/- mice (AChE knockout mice) of various ages.METHODS:Cultured ARPE-19 cells were treated with hydrogen peroxide (H2O2) at concentrations of 0, 250, 500, 1000 and 2000 μmol/L and protein levels were measured using Western blot. Intraperitoneal injections of tacrine and donepezil (0.1 mg/mL, 0.2 mg/mL and 0.4 mg/mL) were respectively given to AChE^+/- mice aged 2mo and 4mo and wild-type S129 mice for 7d; phosphate buffered saline (PBS) was administered to the control group. The mice were sacrificed after 30d by in vitro cardiac perfusion and retinal samples were taken. AChE-deficient mice were identified by polymerase chain reaction (PCR) analysis using specific genotyping protocols obtained from the Jackson Laboratory website. H&E staining, immunofluorescence and Western blot were performed to observe AChE protein expression changes in the retinal pigment epithelial (RPE) cell layer.RESULTS:Different concentrations of H2O2 induced AChE expression during RPE cell apoptosis. AChE^+/- mice retina were thinner than those in wild-type mice (P<0.05); the retinal structure was still intact at 2mo but became thinner with increasing age (P<0.05); furthermore, AChE^+/- mice developed more slowly than wild-type mice (P<0.05). Increased concentrations of tacrine and donepezil did not significantly improve the protection of the retina function and morphology (P>0.05).CONCLUSION:In vivo, tacrine and donepezil can inhibit the expression of AChE; the decrease of AChE expression in the retina is beneficial for the development of the retina.     

A new computational model for human thyroid cancer enhances the preoperative diagnostic efficacy

Considering the high rate of missed diagnosis and delayed treatments for thyroid cancer, an effective systematic model for the differential diagnosis is highly needed. Thus we analyzed the data on the clinicopathological characteristics, routine laboratory tests and imaging examinations in a cohort of 13,980 patients with thyroid cancer to establish a new diagnostic model for differentiating thyroid cancer in clinical practice. Here, we randomly selected two-thirds of the population to develop the thyroid malignancy risk scoring system (TMRS) for preoperative differentiation between thyroid cancer and benignant thyroid diseases, and then validated its differential diagnostic power in the rest one-third population. The 18 predictors finally enrolled in the TMRS included male gender, clinical manifestations (fever, neck sore, neck lump, palpitations or sweating), laboratory findings (TSH>1.56mIU/L, FT₃>5.85pmol/L, TPOAb>14.97IU/ml, TgAb>48.00IU/ml, Tg>34.59μg/L, Ct>64.00ng/L, and CEA>0.41μg/L), and ultrasound features (tumor number≤ 23mm, site, size, echo texture, margins, and shape of neck lymphnodes). The TMRS is validated to be well-calibrated (P = 0.437) and excellently discriminated (AUC = 0.93, 95% CI [0.92, 0.94]), with an accuracy of 83.2%, a sensitivity of 89.3%, a specificity of 81.5%, positive and negative predictive values of 56.8% and 96.6%, positive and negative likelihood ratios of 4.83 and 0.13 in the development cohort, respectively. The TMRS highlights that this differential diagnostic system could help provide accurate preoperative risk stratification for thyroid cancer, and avoid unnecessary over- and under-treatment for such patients.     

Chemokines CCL2, 3, 14 stimulate macrophage bone marrow homing,proliferation, and polarization in multiple myeloma

We previously showed that macrophages (MΦs) infiltrate the bone marrow (BM) of patients with myeloma and may play a role in drug resistance. This study analyzed chemokines expressed by myeloma BM that are responsible for recruiting monocytes to the tumor bed. We found that chemokines CCL3, CCL14, and CCL2 were highly expressed by myeloma and BM cells, and the levels of CCL14 and CCL3 in myeloma BM positively correlated with the percentage of BM-infiltrating MΦs. In vitro, these chemokines were responsible for chemoattracting human monocytes to tumor sites and in vivo for MΦ infiltration into myeloma-bearing BM in the 5TGM1 mouse model. Surprisingly, we also found that these chemokines stimulated MΦ in vitro proliferation induced by myeloma cells and in vivo in a human myeloma xenograft SCID mouse model. The chemokines also activated normal MΦ polarization and differentiation into myeloma-associated MΦs. Western blot analysis revealed that these chemokines promoted growth and survival signaling in MΦs via activating the PI3K/Akt and ERK MAPK pathways and c-myc expression. Thus, this study provides novel insight into the mechanism of MΦ infiltration of BM and also potential targets for improving the efficacy of chemotherapy in myeloma.     

High preoperative serum globulin in rectal cancer treated with neoadjunctive chemoradiation therapy is a risk factor for poor outcome

An elevated serum albumin (ALB) and albumin/globulin ratio (AGR) has been reported to be associated with a favorable prognosis for certain malignancies; however, little is known about the prognostic significance of globulin (GLB) in rectal cancer treated with neoadjuvant chemoradiation therapy (NCRT). The purpose of this study was to evaluate whether GLB analysis could predict the prognosis of patients received NCRT. A retrospective cohort of 293 locally advanced rectal cancer patients receiving NCRT followed by radical surgery was recruited between January 2006 and December 2012 at Fudan University Shanghai Cancer Center. Levels for preoperative GLB and ALB were obtained and used to calculate the AGR. Survival analysis was used to evaluate the predictive value of GLB. X-tile program determined 28.50, 36.20 and 1.20 as optimal cut-off value for GLB, ALB and AGR in terms of survival. Univariate and multivariate analysis revealed that low GLB levels were significantly associated with favorable rectal cancer-specific survival (RCSS) (P < 0.05). Conversely, low ALB levels were associated with a significantly worse RCSS (P = 0.010). Collectively, high preoperative GLB level was a significantly unfavorable factor for rectal cancer patients treated with NCRT. This easily obtained variable may serve as a valuable marker to predict the outcomes of such patient population.     

大鼠脑内囊出血模型的构建及其评价

背景:稳定准确的动物模型是研究出血性脑血管病的必要工具和基础。目的:建立和评价大鼠脑内囊出血模型。设计:随机对照动物实验。单位:徐州医学院第二附属医院;南京医科大学第一附属医院。材料:实验于2002-05/11在南京医科大学动物实验中心完成。35只SD大鼠随机分为两组:实验组30只,假手术组5只。方法:①通过立体定向术向实验组大鼠脑内囊注入自体血制成脑内囊出血模型。②按ZeaLonga5分制神经病学评分标准评分,观察大鼠躯体感觉及运动功能。③大鼠在麻醉状态下于术前及术后检测体感诱发电位。④测定体感诱发电位后,将大鼠麻醉后处死,取脑制备切片,苏木精-伊红染色,以最大病灶处光镜观察血肿及组织形态学的改变。主要观察指标:①两组大鼠神经功能评分。②两组大鼠体感诱发电位各波潜伏期。③两组大鼠脑组织形态学观察。结果:35只大鼠均进入结果分析。①以出现明显偏瘫为造模成功,本实验成功率为93.3%(28/30),实验组神经病学评分为(2.74±0.46)分,与假手术组(0分)比较,差异有显著性(P<0.05)。②体感诱发电位显示,实验组术后各波潜伏期较术前和假手术组明显延迟[P1:(15.72±0.78)ms,(10.69±0.52)ms,(10.73±0.48)ms;N1:(17.95±1.27)ms,(13.21±1.31)ms,(13.34±1.27)ms;N2:(21.16±1.62)ms,(15.42±1.46)ms,(15.58±1.44)ms;N3:(24.86±1.58)ms,(18.72±1.76)ms,(18.99±1.67)ms,P<0.05]。③假手术组仅见针道周围散在红细胞,无出血灶;实验组病理形态学表现为左侧内囊区不规则或椭圆形血凝块,大致有一个低倍视野范围,出血灶边缘脑组织疏松水肿,病变明显重于假手术组。结论:用立体定向术回注自体血制成大鼠脑内囊出血模型更接近临床脑出血,且方法简便,重复性好。     

Characterization of neuroblastoma bone invasion/metastasis in established bone metastatic model of SY5Y and KCNR cell lines

Objects

To determine the mechanism of neuroblastoma (NB) bone invasion/metastasis, it is necessary to investigate the bone invasion/metastasis-related factors in the bone invasion/metastasis process. Some evidence has suggested that various proteins were involved in bone osteolytic response. The invasion/metastasis property and gene expression of NB, however, are still unknown.

Methods

Single-cell suspensions of SY5Y and KCNR cells were injected directly into the femur of nude mice. Radiological and histological analyses, immunohistochemistry analyses, and western blot assay were performed to characterize bone metastasis mechanism in these bone metastasis models.

Results

SY5Y and KCNR NB cells result in osteolytic responses in bone metastasis model. Osteoprotegerin (OPG), receptor activator of NF-kappaB ligand (RANKL), parathyroid hormone-related peptide (PTHrP), endothelin 1 (ET-1), and CXCR4 were examined and compared among in vitro, in vivo, and normal bone, respectively. PTHrP, OPG, RANKL, and ET-1 except CXCR4 in SY5Y and KCNR NB cells xenografts were strikingly upregulated compared with normal bone and NB cells. However, significantly stronger expression of PTHrP and RANKL was presented than ET-1 and OPG; furthermore, the ratios of expression of PTHrP, RANKL to OPG, and ET-1 were also markedly increased in vivo versus in vitro.

Conclusions

Our study provided evidence that NB cell may enhance bone invasion through PTHrP, OPG, RANKL, and ET-1, especially PTHrP and RANKL which may display stronger effects. CXCR4 appeared not participating in bone invasion, but in tumor growth, and homing to bone. Targeting PTHrP, OPG, ET-1, and RANKL may provide a new insight and method for patient therapy by inhibiting NB bone metastasis and invasiveness.     

Amyloid plaque pathogenesis in 5XFAD mouse spinal cord: retrograde transneuronal modulation after peripheral nerve injury

The spinal cord is composed of distinct neuronal groups with well-defined anatomic connections. In some transgenic (Tg) models of Alzheimer’s disease (AD), amyloid plaques develop in this structure, although the underlying cellular mechanism remains elusive. We attempted to explore the origin, evolution, and modulation of spinal β-amyloid (Aβ) deposition using Tg mice harboring five familiar AD-related mutations (5XFAD) as an experiential model. Dystrophic neuritic elements with enhanced β-secretase-1 (BACE1) immunoreactivity (IR) appeared as early as 2 months of age, and increased with age up to 12 months examined in this study, mostly over the ventral horn (VH). Extracellular Aβ IR emerged and developed during this same period, site-specifically co-existing with BACE1-labeled neurites often in the vicinity of large VH neurons that expressed the mutant human APP. The BACE1-labeled neurites almost invariably colocalized with β-amyloid precursor protein (APP) and synaptophysin, and frequently with the vesicular glutamate transporter-1 (VGLUT). Reduced IR for the neuronal-specific nuclear antigen (NeuN) occurred in the VH by 12 months of age. In 8-month-old animals surviving 6 months after a unilateral sciatic nerve transection, there were significant increases of Aβ, BACE1, and VGLUT IR in the VN of the ipsilateral relative to contralateral lumbar spinal segments. These results suggest that extracellular Aβ deposition in 5XFAD mouse spinal cord relates to a progressive and amyloidogenic synaptic pathology largely involving presynaptic axon terminals from projection neurons in the brain. Spinal neuritic plaque formation is enhanced after peripheral axotomy, suggesting a retrograde transneuronal modulation on pathogenesis.     

Increased dopamine receptor signaling and dopamine receptor-G protein coupling in denervated striatum

Chronic interruption of the nigrostriatal dopaminergic pathway leads to sensitized dopaminergic responses in striatum. We attempted to explore the mechanism(s) underlying this dopaminergic supersensitivity by assessing dopamine receptor signaling and receptor-G protein coupling in unilateral 6-hydroxydopamine-lesioned rats. Dopamine-stimulated adenylyl cyclase activity as well as dopamine-activated guanosine 5'-O-(3-[(35)S]thiotriphosphate) ([(35)S]GTPgammaS) binding and [(3)H]palmitate incorporation by Galpha proteins were enhanced in tissues obtained from denervated striata without apparent changes in Galpha protein levels. Moreover, high-affinity binding sites of the D(1) dopamine receptor increased in lesioned compared with control striata without altering the expression level of the receptor. These denervation-mediated changes appear to correlate with the increase in D(1) dopamine receptor binding sites that co-immunoprecipitated with Galphas(olf)/q(11) proteins. In contrast, the total number of D(2) receptor binding sites was increased, yielding an increase in absolute number of high-affinity sites without significant changes in the proportion of high-affinity sites. Stimulation of the D(2) dopamine receptor enhanced coupling to Galphai protein; this was increased in the striata lesioned. The results provide an important molecular mechanism by which dopamine receptor-regulated signaling is enhanced following denervation of dopaminergic input to striatum. Although D(1) dopamine receptor supersensitivity appears to be mediated by enhanced coupling of the receptor to its G proteins, sensitization in the D(2) dopamine receptor system is mediated by increased D(2) receptor density and enhanced D(2) receptor-Gi protein coupling.