肺炎衣原体对C57BL/6J小鼠腹膜巨噬细胞泡沫化的影响

目的:探讨肺炎衣原体在C57BL/6J小鼠腹膜巨噬细胞泡沫化过程中的作用。方法:将肺炎衣原体与C57BL/6J小鼠腹膜巨噬细胞培养72h后用油红O方法染色观察细胞的形态变化并测定细胞内胆固醇的含量。结果:肺炎衣原体能使C57BL/6J小鼠腹膜巨噬细胞胞浆内脂质颗粒增多,胆固醇酯占总胆固醇的比例增加。结论:肺炎衣原体能促进C57BL/6J小鼠腹膜巨噬细胞泡沫化。     

C57BL/6J小鼠主要器官的组织学数据库的建立

目的建立C57BL/6J小鼠主要器官的组织学数据库。方法选取C57BL/6J小鼠雌性10只,雄性10只,将其麻醉致死,解剖切取心、肝、脾、肺、肾、胃等45个器官和组织,用10%的中性缓冲福尔马林固定,石蜡切片,常规HE染色,用显微镜观察,图像采集,用计算机对图像进行标注。采用SQLSEVER2000数据库管理系统,C#编程语言,B/S（浏览器/服务器）为系统构架。结果解剖20只C57BL/6J小鼠主要器官和组织,制做了1 589张切片,观察采集了13 578张图片,对1 563张图片进行了标注,建立了容量达7.85 GB的C57BL/6J小鼠主要器官的组织学数据库。结论构建可查询的C57BL/6J小鼠主要器官的形态学数据库,是研究疾病模型小鼠方便快捷的对照图库。     

一种新型荧光(mKate2)特异性标记中性粒细胞的转基因小鼠的构建及初步应用

目的构建在中性粒细胞中特异性表达mKate2红色荧光蛋白的转基因小鼠。方法本研究将人工合成的lysozyme M启动子及mKate2基因序列构建至pmini Tol2系统中,通过显微注射法把线性化的转基因载体注射到C57BL/6J小鼠的受精卵后并将其移植至假孕鼠体内,制备转基因小鼠。本研究使用PCR鉴定lyz-m Kate2转基因小鼠的成功构建和通过荧光显微镜和流式分析等方法鉴定该模型小鼠中荧光蛋白mKate2的表达及特异性。结果外源lyz-mKate2转基因盒在C57BL/6J小鼠体内成功表达。在外周血中检测到标记红色荧光信号的中性粒细胞,而且70%左右的mKate2阳性细胞是中性粒细胞。激光诱导血栓模型中血栓块可观察到mKate2阳性细胞。结论本研究成功构建了mKate2特异性标记中性粒细胞的新型转基因小鼠并验证了其在血栓模型中的应用价值。     

CXC趋化因子受体2(CXCR2)参与脓毒症脑病小鼠脑内皮细胞激活并介导中性粒细胞迁移

目的探讨CXC趋化因子受体2(CXCR2)在脂多糖(LPS)诱导的脓毒症脑病中对脑内皮细胞激活和中性粒细胞迁移入脑的影响。方法设置C57BL/6J小鼠和CXCR2基因敲除小鼠正常对照组、野生型小鼠LPS处理组和CXCR2基因敲除小鼠LPS处理组。腹腔注射LPS建立脓毒症脑病模型,醋酸AS-D萘酚酯酶组织化学染色检测小鼠脑皮质区域内中性粒细胞的浸润情况, ELISA检测小鼠全脑和血浆中肿瘤坏死因子α(TNF-α)和CXC趋化因子配体1(CXCL1)的水平。分离脑皮质部位微血管内皮细胞并进行原代培养,Western blot法检测LPS(1μg/mL)和TNF-α(200 ng/mL)对原代小鼠脑微血管脑内皮细胞CXCR2蛋白表达的影响以及CXCL1对脑内皮细胞骨架蛋白纤维型肌动蛋白(F-actin)及血管细胞黏附分子1(VCAM-1)蛋白表达的影响。结果腹腔注射LPS后,野生型小鼠(C57BL/6J小鼠)脑和血浆中TNF-α水平升高,同时脑中CXCL1水平也显著升高,且小鼠脑皮质内浸润的中性粒细胞数目显著增多;然而CXCR2基因敲除小鼠脑皮质内浸润的中性粒细胞数目较野生型小鼠显著减少。体外使用LPS和TNF-α刺激后,脑内皮细胞CXCR2蛋白水平升高; CXCL1刺激脑内皮细胞后, F-actin和VCAM-1的蛋白水平也明显升高。结论 CXCR2参与脓毒症脑病小鼠脑内皮细胞的激活,进而介导中性粒细胞的迁移。     

γ射线对C57BL/6J和C3H/HeN小鼠肺成纤维细胞生物学行为影响的比较研究

目的比较C57BL/6J和C3H/HeN两种小鼠肺成纤维细胞以不同剂量的60Coγ射线照射后生物学行为的异同。方法原代分离培养C57BL/6J和C3H/HeN两种小鼠肺成纤维细胞(LF),应用2、4、6和8Gy的60Coγ射线照射后,通过MTT比色法、流式细胞术、AgNOR染色和免疫荧光细胞化学染色法,检测照射后两种LF的增殖活力、细胞周期以及a-平滑肌肌动蛋白(a-SMA)、基质金属蛋白酶-1(MMP-1)和金属蛋白酶组织抑制剂-1(TIMP-1)表达的变化。结果以2~8Gy照射后,C57BL/6J和C3H/HeN小鼠的LF增殖活力与正常对照组相比较未见明显增强。照射后,C57BL/6J小鼠的LF非整倍体增多,a-SMA高表达,MMP-1的表达呈弱阳性,TIMP-1的表达呈增强趋势。照射后,C3H/HeN小鼠的LF出现G2-M期阻滞,a-SMA的表达减弱至消失,MMP-1和TIMP-1的表达均呈增强趋势。结论以60Coγ射线照射后,C57BL/6J和C3H/HeN两种小鼠的LF生物学行为不同,C57BL/6J小鼠的LF呈“活化”状态,为该种小鼠易发生肺纤维化的细胞学基础提供了实验依据。     

饥饿、糖尿病和肥胖状态对小鼠肝脏SOCS2基因表达的影响

目的确定小鼠肝脏SOCS2基因在饥饿、糖尿病和肥胖状态下的表达水平,并初步研究SOCS2对糖异生的影响。方法动物分3组:C57BL/6J小鼠、对照组(饱食)和实验组(饥饿24 h);糖尿病模型小鼠db/db及对照小鼠db/m饱食;肥胖模型小鼠ob/ob及其对照C57BL/6J小鼠(饱食)。处死小鼠后提取肝脏RNA做反转录PCR,荧光实时定量PCR检测小鼠肝脏SOCS2及糖异生相关基因在3组小鼠中的表达水平;使用腺病毒表达系统在C57BL/6J小鼠原代肝细胞中过表达SOCS2,Western blot检测SOCS2蛋白的表达,葡萄糖生成实验检测糖输出。结果饥饿使C57BL/6J小鼠肝脏中SOCS2 mRNA水平下调,db/db和ob/ob小鼠肝脏SOCS2基因表达比其对照小鼠均明显下降(P0.05),调节糖异生的关键基因PGC-1α、PEPCK和G6Pase的mRNA水平均上升。在C57BL/6J小鼠原代肝细胞中过表达SOCS2,得到大小为Mr 23 000的蛋白,糖输出受到明显抑制。结论初步认定SOCS2可抑制C57BL/6J小鼠原代肝细胞糖异生,可能是治疗糖尿病的一个新靶点。     

Api6在高脂高胆固醇饮食引起肺部炎症中所发挥作用的研究*

 目的：研究凋亡抑制因子6（Api6）在高脂高胆固醇饮食所致C57BL/6J小鼠肺部炎症反应中的作用。方法：6～8周龄的C57BL/6J雄性小鼠喂养于SPF环境中,随机分成2组,分别给予普通饮食和高脂高胆固醇饮食喂养。喂养16周后收集肺组织并采用免疫组织化学和ELISA法鉴定肺组织的炎症状态。实时定量PCR和Western blotting鉴定Api6 mRNA与蛋白的表达水平,流式细胞术检测小鼠支气管肺泡灌洗液细胞凋亡情况。体外培养巨噬细胞RAW264.7,流式细胞术检测Api6对氧化型低密度脂蛋白（oxLDL）引起的细胞凋亡的影响。结果：高脂高胆固醇饮食喂养小鼠16周后,C57BL/6J小鼠肺组织出现以巨噬细胞蓄积以及肿瘤坏死因子α和单核细胞趋化蛋白1升高为主的炎症反应。与普通饮食组相比,高脂高胆固醇饮食喂养小鼠肺组织的Api6 mRNA和蛋白表达水平都显著上调（P<0.01）,同时支气管肺泡灌洗液中的巨噬细胞凋亡水平明显下降（P<0.01）。体外实验证实500 μg/L的重组Api6处理RAW264.7细胞可显著抑制oxLDL引起的细胞凋亡（P<0.05）。结论：高脂高胆固醇饮食可致C57BL/6J小鼠肺组织巨噬细胞蓄积,其机制可能与Api6抑制巨噬细胞的凋亡有关。     

miR-200C与miR-16在哮喘患儿血清及哮喘小鼠肺组织中的表达上调

目的探讨miR-200C与miR-16在支气管哮喘患儿外周血淋巴细胞及支气管哮喘小鼠模型肺组织中的表达。方法应用real-time PCR方法检测支气管哮喘患儿与正常儿童外周血淋巴细胞miR-200C与miR-16的表达,卵蛋白致敏方法建立支气管哮喘小鼠模型,肺泡灌洗液(BALF)细胞计数,HE染色观察气道炎症。应用real-time PCR法检测哮喘组小鼠与对照组小鼠肺组织miR-200C与miR-16的表达。结果与正常儿童相比,支气管哮喘患儿外周血淋巴细胞miR-200C与miR-16表达水平显著升高(P0.01)。哮喘小鼠模型组BALF中细胞总数及嗜酸性粒细胞数,炎症细胞浸润,肺组织miR-200C与miR-16表达水平显著高于正常对照组(P0.01)。结论miR-200C与miR-16在支气管哮喘患儿及支气管哮喘小鼠模型中高表达。     

肺炎衣原体感染致C57BL/6J小鼠内皮功能异常

目的：研究肺炎衣原体感染对C57BL/6J小鼠主动脉内皮依赖舒张反应及动脉粥样硬化形成的影响。方法：32只C57BL/6J小鼠分为肺炎衣原体感染并胆固醇饲料喂养组、胆固醇饲料喂养组、肺炎衣原体感染组和对照组，喂养24周，取主动脉弓标本分析动脉粥样硬化斑块面积，远端胸主动脉作血管舒张功能测定，血样品作血脂、一氧化氮水平及内皮素浓度测定。结果：肺炎衣原体感染并胆固醇饲料喂养组、胆固醇饲料喂养组和肺炎衣原体感染组乙酰胆碱引起的动脉平均最大舒张百分数明显低于对照组(P＜0.01)，且一氧化氮水平也较低，单纯肺炎衣原体感染小鼠无明显动脉粥样硬化形成。结论：肺炎衣原体感染可损害C57BL/6J小鼠动脉内皮功能，一氧化氮途径可能参与其发展。     

炎症反应通过上调CD36表达促进小鼠肾脏损伤

目的:探讨慢性炎症对小鼠肾脏CD36表达的影响及其在小鼠肾脏损伤中的作用。方法:将8周龄雄性C57BL/6J小鼠和CD36基因敲除(CD36KO)小鼠随机分为C57BL/6J生理盐水注射组、C57BL/6J酪蛋白注射组和CD36KO酪蛋白注射组,每组8只。高脂喂养处理14周后,收集小鼠血清、24 h尿液和肾组织样本。ELISA试剂盒检测血清中肿瘤坏死因子α(TNF-α)含量,全自动生化仪测定血、尿肾功能指标,HE染色和Masson染色分析肾脏病理改变,real-time PCR和Western blot检测肾脏组织中CD36及炎症/趋化因子(MCP-1、IL-6和TNF-α)m RNA和蛋白的表达,试剂盒测定组织内过氧化氢含量,免疫组化染色测定肾组织Nrf2和TGF-β1的蛋白表达。结果:与生理盐水注射组相比,酪蛋白注射能增强C57BL/6J小鼠血清TNF-α含量和肾组织中TNF-α的蛋白表达(P 0. 05),提示酪蛋白注射能成功诱导小鼠全身和肾脏局部的慢性炎症。同时酪蛋白注射显著促进了小鼠肾组织的CD36和TGF-β1蛋白表达,引起肾小球硬化、蛋白尿和血清肌酐含量显著增加,组织过氧化氢含量明显增加,Nrf2含量和抗氧化能力明显降低(P 0. 05)。而酪蛋白处理的CD36基因敲除小鼠肾组织病理学改变不明显,血、尿肾功能指标和尿量较酪蛋白处理的C57BL/6J小鼠明显降低,且肾组织过氧化氢含量低于酪蛋白处理的C57BL/6J小鼠(P 0. 05)。结论:炎症应激通过促进小鼠肾组织CD36表达,促进氧化应激,导致小鼠肾损伤。     

The relative difference in anti-Listeria resistance of C57BL/6 and A/J mice is not eliminated by active immunization or by transfer of Listeria-immune T cells.

In this study, we examined the effects of active and adoptive immunization on the anti-Listeria resistance of innately resistant C57BL/6 and innately susceptible A/J mice. Although active immunization with a sublethal dose of viable Listeria monocytogenes markedly enhanced the anti-Listeria resistance of both C57BL/6 and A/J mice, the 100-fold difference between the two strains in innate anti-Listeria resistance was not diminished. Following immunization with an equivalent sublethal dose (0.1 LD50) of L. monocytogenes, both C57BL/6 and A/J mice generated T cells that could transfer significant and comparable protection to syngeneic recipients that were challenged with up to a 10 LD50 dose of L. monocytogenes. When the absolute number of viable Listeria was compared, however, it was clear that T cells from immunized C57BL/6 mice were capable of transferring protection to syngeneic recipients at Listeria challenge doses that were more than 100-fold greater than could T cells from Listeria-immunized A/J mice. Both active immunization and adoptive transfer of syngeneic Listeria-immune T cells enhanced the accumulation of inflammatory neutrophils and macrophages in C57BL/6 and A/J mice. More inflammatory neutrophils were recovered from actively immunized C57BL/6 than from A/J mice, whereas more inflammatory macrophages were obtained from adoptively immunized C57BL/6 than from A/J mice. These results provide further evidence for the beneficial role of inflammation in genetically determined innate resistance and T-cell mediated resistance to listeriosis. These data also suggest that some mechanism in addition to inflammatory responsiveness may be responsible for limiting the expression of acquired anti-Listeria resistance in genetically susceptible A/J mice.     

Genetically determined resistance to listeriosis is associated with increased accumulation of inflammatory neutrophils and macrophages which have enhanced listericidal activity.

The C57BL/6 and A/J inbred strains of mice differ markedly in their resistance to the facultative intracellular bacterium Listeria monocytogenes. One possible explanation for this genetically determined resistance is that phagocytes from Listeria-resistant strains of mice can kill L. monocytogenes more effectively than phagocytes from Listeria-susceptible strains of mice. We report here that inflammatory neutrophils and macrophages from Listeria-resistant mice (C57BL/6) exhibit a slight but significantly enhanced ability to kill L. monocytogenes in vitro as compared to inflammatory phagocytes from Listeria-susceptible mice (A/J). More importantly, however, Listeria-resistant mice recruited more inflammatory neutrophils and macrophages to the peritoneal cavity in response to i.p. injection of heat-killed Listeria than did Listeria-susceptible mice. These data suggest that genetically determined resistance to listeriosis is dependent on the enhanced inflammatory responsiveness of Listeria-resistant mice. Further support for this hypothesis was provided by experiments in which the passive transfer to A/J mice (C5-deficient) of plasma from C57BL/6 mice (C5-sufficient) enhanced the ability of the recipient A/J mice both to recruit inflammatory neutrophils to the peritoneal cavity in response to i.p. injection of heat-killed Listeria, and to clear L. monocytogenes from the spleen after a sublethal challenge of viable Listeria.     

Mice: progesterone and the regulation of strain differences in pregnancy-induced nest building

Pregnant DBA/2J females built significantly larger and more completely enclosed nests than did pregnant C57BL/6J mice. This strain difference was restricted to the last half of gestation and was not observed during either the virgin state or lactation. Genotype-based differences in pregnancy-induced nest building were not related to circulating levels of progesterone (P), core temperature, or body weight. Exposure to supplemented P during pregnancy elevated nest building exhibited by pregnant C57BL females but did not induce DBA-like levels of the behavior. Also, virgin DBA females built larger nests in response to P than did C57BL females. These findings suggest that differences in the sensitivity of central neural tissue to steroid hormones may account for genotypically determined variation in patterns of pregnancy-induced nest building.     

Murine MicroRNA‐214 regulates intracellular adhesion molecule (ICAM1) gene expression in genital Chlamydia muridarum infection

The hallmark of chlamydial infection is the development of upper genital pathology in the form of hydrosalpinx and oviduct and/or tubal dilatation. Although molecular events leading to genital tissue presentation and cellular architectural remodelling are unclear, early-stage host immune responses are believed to contribute to these long-term sequelae. Recently, we reported the contribution of selected infection-associated microRNAs (miRs) in the generation of host immunity at early-stage infection (day 6 after intravaginal Chlamydia muridarum challenge in C57BL/6 mice). In this report, we describe the contribution of an infection-associated microRNA, i.e. miR-214, to host immunity. Chlamydia muridarum infection in the C57BL/6 mouse genital tract significantly down-regulated miR-214 while up-regulating intracellular adhesion molecule 1 (ICAM1) gene expression. These in vivo observations were confirmed by establishing direct regulation of ICAM-1 by miR-214 in ex vivo genital cell cultures in the presence of miR-214 mimic and inhibitor. Because, ICAM-1 contributes to recruitment of neutrophils following infection, we also demonstrated that alteration of ICAM1 by miR-214 in interleukin-17A-deficient (IL-17A^−/−) mice correlated with reduction of neutrophils infiltrating genital tissue at day 6 after challenge. Additionally, these early-stage events resulted in significantly decreased genital pathology in IL-17A^−/− mice compared with C57BL/6 mice. This report provides evidence for early-stage regulation of ICAM1 by microRNAs, resulting in reduction of genital pathology associated with chlamydial infection.     

Intranasal infection of beige mice with Mycobacterium avium complex: role of neutrophils and natural killer cells.

Beige mice show increased susceptibility to intranasal infection with organisms of the Mycobacterium avium complex (MAC) compared with their immunocompetent congenics, C57BL/6 mice. This increased susceptibility was clear 2 weeks postinfection, before the activation of the specific immune response. T lymphocytes from 4-week infected beige mice, cultured in vitro, produced amounts of gamma interferon similar to those found in cells from C57BL/6 mice. Macrophage activation, as judged by NO production and lysis of the macrophage target P815, occurred in the lungs of beige mice. Despite the inability of bone marrow-derived NK cells from beige mice to lyse NK-susceptible YAC-1 cells, their gamma interferon production was normal. Monoclonal antibody to NK1.1 was used to deplete C57BL/10 mice of lytic activity against YAC-1 cells without exacerbating infection between 2 and 6 weeks of observation, making it unlikely that any deficiency in NK cells was the cause of susceptibility in beige mice. There was a striking influx of neutrophils in the lungs of beige mice compared with C57BL/6. More than half of the MAC organisms appeared associated with the neutrophils of beige mice, while in C57BL/6 mice, most MAC organisms were associated with cells of macrophage/monocyte morphology. Injection of monoclonal antibody specific for neutrophils failed to eliminate those cells from the lungs of beige mice. However, in C57BL/6 mice, neutrophil numbers were reduced by 95% without exacerbating the infection. We conclude that, although neutrophils are not essential to the relative resistance of C57BL/6 mice, the known deficiencies in both neutrophils and macrophages account for the susceptibility of beige mice.     

LPS binding protein is important in the airway response to inhaled endotoxin

BACKGROUND: Inhaled endotoxin is a risk factor for asthma exacerbation, and endotoxin inhalation by itself recapitulates many of the classical features of asthma in mice, including reversible airflow obstruction and inflammation, airways hyperresponsiveness, and airway remodeling. OBJECTIVE: Our objective was to determine the importance of LPS binding protein (LBP) in the response to inhaled LPS. METHODS: We challenged LBP-deficient mice (C57BL/6(LBP-/-)) and C57BL/6 mice with inhaled endotoxin for 4 hours, 5 days, or 4 weeks, followed by 3 days of recovery. RESULTS: LBP in the lung was significantly increased in LPS-exposed C57BL/6 mice from all 3 groups. Only LPS-exposed C57BL/6 mice had significantly enhanced airway responsiveness to inhaled methacholine. Total lavage cells in LPS-exposed C57BL/6(LBP-/-) mice were significantly reduced compared with those seen in LPS-exposed C57BL/6 mice; however, the percentage of PMNs was similarly increased in both the C57BL/6 and C57BL/6(LBP-/-) mice. TNF-alpha, IL-1 beta, and IL-6 protein concentrations in whole-lung lavage fluid from C57BL/6(LBP-/-) mice were also significantly reduced when compared with those seen in C57BL/6 mice. In C57BL/6(LBP-/-) mice submucosal cell proliferation was significantly reduced in the 1-week group when compared with that seen in similarly exposed C57BL/6 mice. The 4-week exposed C57BL/6 mice had significantly thickened airway submucosa and significantly increased lavaged TGF-beta(1) protein compared with that seen in C57BL/6(LBP-/-) mice. CONCLUSIONS: These findings indicate that LBP is one of the critical molecules regulating the acute and chronic airway response to inhaled LPS.     

Infanticide: Genetic,developmental and hormonal influences in mice

DBA/2J and androgen-deficient C57BL/6J mice were examined for their response to newborn (1–3 day old) Rockland Swiss (R-S) albino mouse pups. Significantly more 70–90 day old C57BL male mice killed young as compared to similarly aged DBA males (80% vs 30% respectively). Adult, 70–90 day old, female mice of both strains typically retrieved newborn young to the nest site. Adult castration significantly reduced infanticide in males of both strains while exposure to testosterone (T)-containing silastic capsules restored it. Treatment of adult ovariectomized female mice of both strains with T-containing capsules significantly elevated the exhibition of pup killing. When tested for infanticide at 25, 35, 45, 55 or 65 days of age, few males of either strain killed young at 25 days of age. However, beginning at 35 days of age, significantly more C57BL males killed young at every age as compared to DBA males. Moreover, C57BL males exhibited an earlier developmental onset of adult-like levels of infanticide than DBA males (45 vs 65 days of age respectively). Finally, older (4.5 to 7 months of age) DBA males exhibited levels of pup-killing identical to that of younger (2 months of age) C57BL males (70%). The findings are discussed in terms of their relationship to other sexually dimophic T-dependent masculine behaviors. The potential importance of infanticide for rodent reproductive strategies and population regulation is also considered.     

Evoked pain behavior and spinal glia activation is dependent on tumor necrosis factor receptor 1 and 2 in a mouse model of bone cancer pain

Bone–cancer-related pain is one of the most disabling factors in patients suffering from primary bone cancer or bone metastases. Recent studies point toward an important role of proinflammatory cytokines, example tumor necrosis factor-α (TNF), for tumor growth and bone–cancer-associated pain. Mechanisms by which TNF, through its receptor subtypes, TNF receptor 1 (TNFR1) and −2 (TNFR2), elicits altered sensation and pain behavior, are still incompletely understood. To look for a potential role of TNF in bone cancer pain, cancer-related pain was analyzed in fibrosarcoma-bearing C57Bl/6J wild type mice after systemic antagonism of TNF. To further clarify the role of TNF receptor (TNFR) in bone-cancer pain, naive and fibrosarcoma-bearing C57Bl/ 6J wild type and transgenic mice with a deficiency of TNFR1 (TNFR1ko), TNFR2 (TNFR2ko), and TNFR1+2 (TNFR1+2ko) were compared regarding cancer-related pain and hyperalgesia, tumor growth, osteoclast activation, and spinal astrogliosis. Systemic antagonism of TNF significantly alleviated tactile hypersensitivity and spontaneous bone–cancer-related pain behavior. Most interestingly, combined deletion of the TNFR1 and TNFR2, but not of either gene alone, almost completely inhibited the development of tactile hypersensitivity, whereas spontaneous pain behavior was transiently increased. Accordingly, spinal astrogliosis was markedly reduced, whereas tumor growth was significantly increased in TNFR1+2ko mice. In contrast, deletion of the TNFR1 or TNFR2 gene alone did not change tumor growth or spinal astrogliosis. Our findings suggest that the combined absence of TNFR1 and TNFR2 is necessary for the attenuation of cancer-related tactile hypersensitivity and concomitant spinal astrogliosis, whereas tumor growth seems to be inhibited by combined TNFR activation. These findings support the hypothesis of cytokine-dependent pain development in cancer pain. Differential targeting of TNFR activation could be an interesting strategy in bone–cancer-related pain conditions.     

Behavioral Differences among C57BL/6 Substrains: Implications for Transgenic and Knockout Studies

Separate breeding colonies of C57BL/6 (“B6”) mice maintained at the Jackson Laboratories (“J”) and NIH (“N”) have led to the emergence of two distinct substrains of C57BL/6 mice: C57BL/6J and C57BL/6N. Molecular genetic studies indicate simple sequence-length polymorphisms, single-nucleotide polymorphisms, and copy-number variants among B6 substrains that may contribute to phenotypic differences. We examined differences in motor coordination, pain sensitivity, and conditional fear in the C57BL/6J strain and three N strains: C57BL/6NCrl (Charles River), C57BL/6NTac (Taconic), and C57BL/6NHsd (Harlan Sprague Dawley). Male C57BL/6J mice demonstrated enhanced motor coordination, as measured by the rotarod assay, markedly enhanced pain sensitivity in two assays of acute thermal nociception (e.g., tail withdrawal and hot plate), and a reduced level of conditional fear. The tail withdrawal result was confirmed in a separate laboratory. We also provide a table reviewing previously reported behavioral differences among various B6 substrains and discuss the significance of environmental differences due to obtaining mice form different vendors. These data may be seen as a potential problem and as a potential opportunity. Great care must be taken when working with mice engineered by using B6 embryonic stem cell lines because control groups, backcrosses, and intercrosses could inadvertently introduce behaviorally significant polymorphic alleles or environmental confounds. On the other hand, deliberate crosses between B6 substrains may provide an opportunity to map polymorphic loci that contribute to variability in a trait on largely homogenous backgrounds, which has the potential to improve mapping resolution and aid in the selection of candidate genes.     

Genetic and pharmacological manipulation of mu opioid receptors in mice reveals a differential effect on behavioral sensitization to cocaine

Cocaine-induced behavioral sensitization is a complex phenomenon involving a number of neuromodulator and neurotransmitter systems. To specifically investigate the role of the micro opioid receptor (MOR) in cocaine-induced behavioral sensitization in mice, both genetic and pharmacological approaches were undertaken. MOR-1 deficient mice of varying backgrounds (C57BL/6J, 129S6, F1 hybrid 129S6xC57BL/6J and 129S6xC57BL/6J) and wild-type C57BL/6J mice exposed continuously to naltrexone, an opioid receptor antagonist, received single daily injections of saline or cocaine for 10 days. All mice received a single cocaine challenge 7 days following the last saline or cocaine injection to test for the expression of sensitization. The locomotor-stimulating and sensitizing effects of cocaine observed in MOR-1 wild-type mice were absent in MOR-1 knockout mice maintained on the mixed 129S6xC57BL/6J background. In contrast, MOR-1 deficient mice developed on a C57BL/6J background showed an accentuated sensitivity to cocaine-induced locomotion. Cocaine's psychomotor activating effects were more pronounced in the MOR-1 C57BL/6J knockouts injected daily with cocaine than in the MOR-1 wild-type mice. Similar locomotor-stimulating and sensitizing effects were found in both F1 hybrid 129S6xC57BL/6J MOR-1 wild-type and MOR-1 knockout mice, while the 129S6 strain showed an overall indifference to cocaine. That is, both the locomotor-stimulating and sensitizing effects of cocaine were absent in both MOR-1 wild-type and MOR-1 knockout mice maintained on the 129S6 background. Lastly, the locomotor-stimulating and sensitizing effects of cocaine were attenuated in C57BL/6J wild-type mice exposed continuously to naltrexone. Collectively, these data support a role for opioidergic involvement in cocaine-influenced behavior in mice. Moreover, MORs appear to differentially modulate a sensitized response to cocaine in different strains of mice as delineated by MOR-1 gene deletion and pharmacological antagonism.