Cannabinoid-induced conditioned place preference in the spontaneously hypertensive rat–an animal model of attention deficit hyperactivity disorder

Rationale  Cannabis preparations are the most widely consumed illicit drugs, and their use typically begins in adolescence. The prevalence of cannabis abuse is higher in patients with attention deficit/hyperactivity disorder (ADHD) than in the general population, yet, knowledge about the motivational properties of cannabinoids in animal models of ADHD are lacking. Objective  To compare the motivational effects of the synthetic cannabinoid agonist WIN55,212-2 (WIN) in adolescent and adult spontaneously hypertensive rats (SHR), a validated animal model of ADHD, and Wistar rats, representing a “normal” genetically heterogeneous population. We also asked whether the effects of WIN depended (1) on the activation of the cerebral subtype of cannabinoid receptors, namely, the CB₁ cannabinoid receptor and (2) on putative changes by WIN in blood pressure. Methods  WIN was tested under an unbiased conditioned place preference (CPP) paradigm. Blood pressure after WIN administration was also monitored in additional groups of rats. Results  In the Wistar rats, WIN produced place aversion only in the adult but not adolescent rats. In contrast, WIN produced CPP in both adolescent and adult SHR rats. The behavioral effects of WIN were CB₁-mediated and not related to blood pressure. Conclusion  The contrasting effects of WIN in Wistar and SHR, and the higher resistance of adolescent rats to the aversive and rewarding effects of WIN in these two strains suggests that both adolescence and the ADHD-like profile exhibited by the SHR strain constitute factors that influence the motivational properties of cannabinoids.     

A PDZ domain recapitulates a unifying mechanism for protein folding

A unifying view has been recently proposed according to which the classical diffusion-collision and nucleation-condensation models may represent two extreme manifestations of an underlying common mechanism for the folding of small globular proteins. We report here the characterization of the folding process of the PDZ domain, a protein that recapitulates the three canonical steps involved in this unifying mechanism, namely: (i) the early formation of a weak nucleus that determines the native-like topology of a large portion of the structure, (ii) a global collapse of the entire polypeptide chain, and (iii) the consolidation of the remaining partially structured regions to achieve the native state conformation. These steps, which are clearly detectable in the PDZ domain investigated here, may be difficult to distinguish experimentally in other proteins, which would thus appear to follow one of the two limiting mechanisms. The analysis of the (un)folding kinetics for other three-state proteins (when available) appears consistent with the predictions ensuing from this unifying mechanism, thus providing a powerful validation of its general nature.     

Immune deficiencies in chronic intestinal pseudo-obstruction

Aim: Chronic intestinal pseudo-obstruction has been associated with urinary disorders, myopathy, and ophthalmoplegia in adults and cholelithiasis in children. We observed a high percentage of total-parenteral-nutrition-dependent patients with pseudo-obstruction and recurrent infections requiring gammaglobulin infusions. Methods: AH records for 23 children with chronic intestinal pseudo-obstruction (10 females and 13 males, mean age 9.8 y ± 4.9 y, range 4–24 y) referred for a nutritional evaluation from 1992 to 1995 were reviewed. Chronic intestinal pseudo-obstruction was diagnosed by clinical, radiographic findings and antroduodenal manometry. Intestinal full-thickness biopsies were performed in seven children. Results: Hypogammaglobulinemia was diagnosed in 18 patients (78%): 16 patients had various immunoglobulin deficiencies and 2 had selective antibody deficiency. Intravenous gammaglobulin was administered in 14 patients. Other medical conditions affecting the children are summarized as follows: autonomic dysfunction in 10 patients (43%), recurrent hypoglycemia in 9 (39%), asthma in 9 (39%), cholecystitis in 7 (30%), low serum carnitine level in 6 (26%), urinary dysfunction in 6 (26%), pancreatitis in 5 (22%), behavioral problems in 5 (22%), myopathy in 2 (9%), idiopathic thrombocytopenia in 2 (8%), velopharyngeal insufficiency in 1 (4%), oculocutaneous albinism in 1 (4%), Pierre-Robin syndrome in 1 (4%), and protein C deficiency in 1 (4%). Munchausen syndrome was suspected in two patients. Conclusions: Chronic intestinal pseudo-obstruction appears to be associated with immune deficiencies. It is unclear if the immune deficiencies, intestinal pseudo-obstruction, and the other medical conditions have a common underlying etiology. Repeated infections may be due to impaired immune function in children with chronic intestinal pseudo-obstruction. We recommend screening for immune deficiencies in children with chronic intestinal pseudo-obstruction.     

Inter-rater agreement and reliability of the COSMIN (COnsensus-based Standards for the selection of health status Measurement Instruments) Checklist

Background  

The COSMIN checklist is a tool for evaluating the methodological quality of studies on measurement properties of health-related patient-reported outcomes. The aim of this study is to determine the inter-rater agreement and reliability of each item score of the COSMIN checklist (n = 114).     

Results of the tension-free vaginal tape procedure for stress incontinence: Patient''s perspective

BACKGROUND: To evaluate the results of the tension-free vaginal tape procedure (TVT) from a patient's perspective. METHODS: Between May 1999 and January 2002, 90 patients underwent a TVT for genuine stress incontinence (GSI) and mixed incontinence. Prior to the procedure, GSI was confirmed by clinical examination and urodynamic studies. Results were then audited from patient notes and the same patients were sent questionnaires to examine results from a patient perspective. RESULTS: Overall response rate to the questionnaire was 70 (77%). The mean age of the patients was 50.4 years (range 31-83 years). Sixty-one patients had spinal anesthesia, seven had general anesthesia and two had local anesthesia. Mean hospital stay was 3.36 days (range 2-14 days) and mean period from the operation to the time of the survey and audit was 16.34 months (range 3-28; SD 6.92). Thirty-nine (56%) of the 70 patients who answered said that the operation had cured their incontinence, 16 (23%) had an improvement in their symptoms, 7 (10%) had worsening of their symptoms and 8 (11%) felt that the operation did not make any difference. The overall success rate according to the patients' perspective was 79%, whereas our audit showed an overall success rate of 86% (77% and 82%, respectively, when we compared only the 66 patients who had both notes and replies available for analysis). CONCLUSION: Although a patient's perception regarding the success of TVT tends to differ from that of a clinician, it was not found to be statistically significant (P = 0.22, McNemar test). The TVT is a very successful operation, but realistic cure rates should be offered to patients.     

Determination of an ensemble of structures representing the intermediate state of the bacterial immunity protein Im7

We present a detailed structural characterization of the intermediate state populated during the folding and unfolding of the bacterial immunity protein Im7. We achieve this result by incorporating a variety of experimental data available for this species in molecular dynamics simulations. First, we define the structure of the exchange-competent intermediate state of Im7 by using equilibrium hydrogen-exchange protection factors. Second, we use this ensemble to predict Phi-values and compare the results with the experimentally determined Phi-values of the kinetic refolding intermediate. Third, we predict chemical-shift measurements and compare them with the measured chemical shifts of a mutational variant of Im7 for which the kinetic folding intermediate is the most stable state populated at equilibrium. Remarkably, we found that the properties of the latter two species are predicted with high accuracy from the exchange-competent intermediate that we determined, suggesting that these three states are characterized by a similar architecture in which helices I, II, and IV are aligned in a native-like, but reorganized, manner. Furthermore, the structural ensemble that we obtained enabled us to rationalize the results of tryptophan fluorescence experiments in the WT protein and a series of mutational variants. The results show that the integration of diverse sets of experimental data at relatively low structural resolution is a powerful approach that can provide insights into the structural organization of this conformationally heterogeneous three-helix intermediate with unprecedented detail and highlight the importance of both native and non-native interactions in stabilizing its structure.     

小鼠皮肤超氧化物歧化酶活性与枸杞多糖的干预

目的:观察枸杞多糖对皮肤胶原代谢和自由基产生的影响,探讨其抗皮肤衰老的作用。方法:实验于2005-06/2006-05在广东医学院整形外科研究所完成。①实验材料:清洁级昆明小鼠60只,月龄2个月,体质量16～24g,雌雄各半。②实验分组:将小鼠随机分为正常对照组、衰老模型组和抗衰老模型组,每组20只。③实验干预:模型组每日用D-半乳糖溶液皮下注射制造衰老模型,用量和时间为80mg/(kg·d)7d,120mg/(kg·d)14d,140mg/(kg·d)14d,180mg/(kg·d)7d。正常对照组每日注射同体积的生理盐水。抗衰老模型组在注射D-半乳糖期间以枸杞多糖灌胃,剂量为20mg/(kg·d),正常对照组和衰老组则以同体积的生理盐水代之灌胃。④实验评估:42d后切取小鼠颈背部皮肤,测定超氧化物歧化酶活力、羟脯氨酸和丙二醛含量。结果:56只小鼠进入结果分析(4只死亡)。①小鼠皮肤超氧化物歧化酶活力:与正常对照组相比,衰老组和抗衰老组小鼠皮肤超氧化物歧化酶活力降低,差异有显著性意义(P<0.01);抗衰老组与衰老模型组比较,超氧化物歧化酶活力增加,差异有显著性意义(P<0.01)。②与正常对照组相比,衰老组和抗衰老组小鼠皮肤羟脯氨酸和丙二醛含量增加,差异有显著性意义(P<0.01);抗衰老组与衰老组比较,羟脯氨酸和丙二醛含量均降低,差异有显著性意义(P<0.01)。结论:枸杞多糖改善皮肤老化的作用与提高小鼠皮肤超氧化物歧化酶活力,降低羟脯氨酸、丙二醛含量,影响胶原代谢有关。     

三七总皂甙对人肝癌细胞缝隙连接细胞间通讯的调节机制

目的：观察三七总皂甙对人肝癌细胞SMMC-7721缝隙连接细胞间通讯、以及缝隙连接蛋白32mRNA和磷酸化细胞外信号调节激酶1，2表达的影响。方法：实验于2005—08／2006—03在江西省分子医学重点实验室完成。①实验操作：SMMC-7721细胞用含100mg／L，200mg／L，400mg／L三七总皂甙的培养液预处理，对照组SMMC-7721细胞不加三七总皂甙。②实验评估：采用划痕标记，染料示踪技术检测缝隙连接细胞间通讯的变化；采用反转录-聚合酶链反应检测缝隙连接蛋白32mRNA水平；采用流式细胞仪检测磷酸化细胞外信号调节激酶1／2的表达。结果：①对照组染料局限于划痕两侧的细胞内，无明显的荧光染料传输，GJIC阴性；三七总皂甙可上调染料传输范围，且随三七总皂甙剂量增高，染料传输范围逐渐增大，最远可达三四层细胞。②反转录-聚合酶链反应结果显示，各组细胞缝隙连接蛋白32mRNA水平差异无显著性意义（P〉0．05）。③流式细胞仪结果显示，对照组SMMC-7721细胞磷酸化细胞外信号调节激酶1／2表达的阳性率较高，伴随三七总皂甙剂量增加磷酸化细胞外信号调节激酶1／2表达的阳性率逐渐下降，呈一定的剂量依赖效应。结论：三七总皂甙可能通过抑制SMMC-7721细胞磷酸化细胞外信号调节激酶1／2的活化减少缝隙连接蛋白32的磷酸化，进而上调SMMC-7721细胞的缝隙连接细胞间通讯功能。     

Heterotypic adherence between human B-lymphoblastic and pre-B- lymphoblastic cells and marrow stromal cells is a biphasic event: integrin very late antigen-4 alpha mediates only the early phase of the heterotypic adhesion

Heterotypic adherence between marrow stromal cells (MSC) and lymphoblastic cells is essential for normal lymphopoiesis and malignant lymphoblastic development. However, the detailed molecular mechanisms by which this heterotypic adherence occurs are poorly understood. The cell-cell interactions between a B-lymphoblastic cell line (UTMB-460) and a pre-B-cell line (NALM-6) with MSC were chosen as models to investigate potential mechanisms and adhesion molecules involved in the apposition between normal and malignant lymphoblastic cells and MSC. A parallel-flow detachment assay (PFDA) and a 51Cr detachment assay, coupled with monoclonal antibody (MoAb) blocking experiments, were used to quantify the attachment of lymphoblastic cells to confluent monolayers of MSC. The apposition between MSC and B-lymphoblastic cells (UTMB-460 cells) was investigated for variable time periods, ranging from 1 minute to 4 hours. Results from the temporal study suggest that the heterotypic adherence of the B-lymphoblastic cells to MSC is a biphasic event and the interactions occur rapidly (< or = 1 minute) after the two cells come into contact. More specifically, the early phase of adherence (< or = 15 minutes) solely involves very late antigen-4 alpha (VLA-4 alpha)/vascular cell adhesion molecule 1 (VCAM- 1) interactions, as evidenced by the nearly complete inhibition (93%) of UTMB-460 cell adherence in the presence of anti-VLA-4 alpha. The late phase (> or = 30 minutes) proceeds despite the continuous presence of anti-VLA-4 alpha. In addition, the late-phase adherence is not affected by MoAbs to LFA-1, CD44, VCAM-1, E-selectin, or L-selectin, which suggests the possible involvement of other adhesion molecules. Adherence of pre-B-lymphoblastic cells (NALM-6) to MSC is also biphasic. Integrin VLA-4 is again a major player in the early phase of pre-B-lymphoblastic cell/MSC interactions. The early phase of adherence may be important in homing of the malignant lymphoblastic cells to the MSC and the late phase in retention of malignant lymphoblastic cells in the bone marrow.     

B-cell growth factor receptor expression and B-cell growth factor response of leukemic B cell precursors and B lineage lymphoid progenitor cells

The purpose of this study was to analyze the expression of B cell growth factor (BCGF) receptors and to elucidate the biologic effects of biochemically purified natural BCGF at the B cell precursor stage of human B lineage lymphoid differentiation. The specific binding of radioiodinated high-mol-wt BCGF (125I-HMW-BCGF) and low-molecular-wt BCGF (125I-LMW-BCGF) to fresh marrow blasts from B cell precursor acute lymphoblastic leukemia (ALL) patients was initially investigated. The estimated number of radioiodinated BCGF molecules bound per blast ranged from undetectable to 24.3 X 10(3) for HMW-BCGF, and from 11.5 X 10(3) to 457.8 X 10(3) for LMW-BCGF. In 3H-TdR incorporation assays, 75% of cases showed a significant response to LMW-BCGF with a median stimulation index of 9.3. By comparison, only 33% of cases showed a significant response to HMW-BCGF with a median stimulation index of 2.4. Subsequently, B cell precursor colony assays were performed to assess and compare the biologic effects of BCGF on leukemic B lineage lymphoid progenitor cells. Among 28 cases studied, 57% responded to both HMW-BCGF and LMW-BCGF, 21% responded only to LMW-BCGF, and the remaining cases showed no proliferative response to either growth factor. The response patterns of virtually pure populations of FACS- sorted leukemic B cell precursors were essentially identical to the proliferative responses of unsorted leukemic B-cell precursors. Synergistic effects between HMW-BCGF and LMW-BCGF were observed in 80% of the cases that responded to both. The numbers of cell-bound radioiodinated BCGF molecules, the stimulation indices, as well as the number of B cell precursor colonies in BCGF-stimulated cultures showed a marked interpatient variation. Patients with structural chromosomal abnormalities (SCAs) involving 12p11-13 or patients with a Philadelphia chromosome showed a greater HMW-BCGF response at the level of leukemic progenitor cells than did other patients (P = .02). The LMW-BCGF response was significantly greater for patients with SCA than for patients without SCA (P = .04). The response of leukemic progenitor cells to HMW-BCGF or LMW-BCGF did not correlate with sex, age, disease status, FAB morphology, WBC at diagnosis, or immunophenotype. To our knowledge, this study represents the first detailed analyses of BCGF receptor expression and BCGF effects in B cell precursor ALL. The data presented provide direct evidence for the expression of functional receptors for both HMW-BCGF and LMW-BCGF in B cell precursor ALL.