Different regulation of p27 and Akt during cardiomyocyte proliferation and hypertrophy

Postnatal cardiomyocytes normally grow by hypertrophy but show a limited proliferate response to certain stimuli. Although the proliferative capacity declines shortly after birth, neonatal cardiomyocytes can grow both by hypertrophy and by proliferation. Therefore, we have used neonatal cardiomyocytes to investigate the molecular differences between hypertrophic and proliferative growth of cardiomyocytes. Stimulation of neonatal cardiomyocytes with angiotensin II mainly induced hypertrophy, whereas PDGF only had a minor effect on the size of the myocytes. In contrast, PDGF induced significant proliferation in the cardiomyocyte cultures whereas angiotensin II treatment only resulted in a small increase in the number of cells. Measurement of cyclin D-dependent kinase specific phosphorylation of pRb by immunohistochemistry showed that, both stimuli activate the G1 phase of the cell cycle. By western blotting we found that PDGF-induced proliferation correlates with activation of Akt, inactivation of GSK-3β and downregulation of the cyclin-dependent kinase inhibitor p27, whereas angiotensin II only had a small effect on Akt, GSK-3β and p27. Our data support the hypothesis that, the hypertrophic and proliferative responses are both activated by G1 cell cycle molecules. The difference between the two responses appears to be that high amounts of p27 are present during hypertrophic growth, whereas proliferation involves downregulation of p27 and GSK-3β activity and upregulation of Akt.     

Prognostic value of the pretreatment albumin:globulin ratio combined with adult comorbidity evaluation 27 and TMN staging in patients with squamous cell carcinoma of the maxillary sinus

The albumin:globulin (A:G) ratio, adult comorbidity evaluation 27 (ACE-27), and TMN staging have been shown to be strong predictive indicators of the survival of patients with many types of tumours. We have investigated the prognostic value of pretreatment based on the A:G ratio combined with TMN staging and ACE-27 in patients with squamous cell carcinoma (SCC) of the maxillary sinus. We studied 196 patients, and the prognostic value was explored by univariate and multivariate Cox’s hazards analysis. Multivariate analyses suggested that pretreatment A:G ratio was independently associated with overall survival (hazard ratio (HR) 1.542, 95% CI 1.219 to 1.991, p = 0.002); disease-specific survival, (HR 1.499, 95% CI 1.197 to 1.842, p = 0.001); and disease-free survival (HR 1.452, 95% CI 1.207 to 1.834, p < 0.001). Additional prognostic factors shown in the survival analyses included ACE-27, pathological T stage, and pathological N stage. Pretreatment A:G ratio combined with ACE-27 and TMN staging were powerful prognostic indicators of outcome in patients with SCC of the maxillary sinus, which has potentially important ramifications for stratification of the disease in the future.     

Design and encapsulation of anticancer dual HSP27 and HER2 inhibitor into low density lipoprotein to target ovarian cancer cells

Tumor cells overexpress low-density lipoprotein (LDL) receptors (LDL-r). Hence, LDL is proposed as a targeting shuttle of anticancer drugs. Therefore, the objective of this study was to synthesize a dual inhibitor of heat shock protein 27 (HSP27) and human epidermal growth factor receptor 2 (HER2) conjugated with cholesterol and encapsulated into LDL for selective targeting of ovarian cancer cells. In the present study, the anticancer agent and its cholesterol conjugate were successfully prepared and characterized physically for color, shape, and melting point. Moreover, the compounds were chemically characterized for ¹H NMR and ¹³C NMR spectra using FTIR and LCMS/MS. Our results revealed that the prepared anticancer agent and its cholesterol conjugate elicited dual HSP27 and HER2 inhibition, as confirmed using western blotting. The anticancer agent (compound D) entered cells and targeted the HSP27 function, thereby reducing HER2 expression. However, a cholesterol-conjugated anticancer agent (compound F) had high cellular uptake and inhibited the growth of SKOV3 cells after encapsulation into LDL. The obtained results concluded that the design of an LDL-encapsulated cholesterol-conjugated HSP27-HER2 dual inhibitor may be a promising approach to realize specific targeted achieve killing of ovarian cancer.     

HLA-B52-positive vasculo-Behçet disease: usefulness of magnetic resonance angiography, ultrasound study, and computed tomographic angiography for the early evaluation of multiarterial lesions

We report a case of HLA-B52-positive Behçet disease accompanied by multiarterial lesions. A 24-year-old woman was suffering from sporadic high fever and recurrent oral and genital ulcers, and laboratory data revealed severe inflammation. A diagnosis of Behçet disease was made. Magnetic resonance angiography, ultrasound study, and computed tomographic angiography demonstrated multiarterial lesions that had caused no symptoms. These noninvasive examinations were extremely useful in evaluating asymptomatic early vascular lesions.     

白细胞介素-27与病毒感染性疾病的研究进展

白细胞介素(IL)-27作为IL-12家族的新成员,是抗原递呈细胞(APC)的早期产物,主要由激活的单核细胞、树突状细胞和巨噬细胞分泌,同时是具有抗炎和促炎双重属性的细胞因子.目前发现IL-27参与多种病毒感染性疾病的发生、发展过程,其有望成为病毒特异性免疫疾病的新的治疗靶点.     

放疗抑制重建术后人工血管新生内膜的实验研究

目的构建膨体聚四氟乙烯(expanded polytetrafluoroethylene,e PTFE)人工血管置换杂种犬肾动脉平面下腹主动脉的血管重建手术模型。探讨总剂量28Gy的体外分割放射治疗影响重建术后人工血管内膜增生的机制。方法健康杂种犬12只,采用数字表法随机均分成放疗组和对照组。均先行肾下腹主动脉段的人工血管置换重建术。放疗组于术后2周后对标记区血管行总剂量28Gy的体外放疗。对照组则不放疗。重建术后5周观察比较两组移植术后的人工血管新生内膜的增生情况,Skp2、p27kip1和增生细胞核抗原(proliferating cell nuclear antigen,PCNA)的蛋白和mRNA的表达情况。结果放疗组的人工血管吻合口内膜增生程度明显比对照组降低(P<0.01)。放疗组的Skp2、PCNA的蛋白表达及mRNA表达较对照组明显减低,而p27kip1的mRNA表达较对照组明显增加(P<0.01)。结论术后体外分割放疗(28Gy)在短期内可以通过抑制血管平滑肌细胞(vascular smooth muscle cell,VSMC)的增生,从而抑制重建术后的人工血管的内膜增生。     

Cyclin-dependent kinase inhibitors p21 and p27 function as critical regulators of liver regeneration following 90% hepatectomy in the rat

BACKGROUNDLiver reduction is the main curative treatment for primary liver cancer, but its use remains limited as liver regeneration requires a minimum of 30% functional parenchyma. AIMTo study the dynamics of the liver regeneration process and consequent behavior of cell cycle regulators in rats after extended hepatectomy (90%) and postoperative glucose infusions.METHODSPost-hepatectomy liver failure was triggered in 84 Wistar rats by reducing their liver mass by 90%. The animals received a post-operative glucose infusion and were randomly assigned to two groups: One to investigate the survival rate and the other for biochemical analyses. Animals that underwent laparotomy or 70% hepatectomy were used as controls. Blood and liver samples were collected on postoperative days 1 to 7. Liver morphology, function, and regeneration were studied with histology, immunohistochemistry, and western blotting. RESULTSPostoperative mortality after major resection reached 20% and 55% in the first 24 h and 48 h, respectively, with an overall total of 70% 7 d after surgery. No apparent signs of apoptotic cell death were detected in the extended hepatectomy rat livers, but hepatocytes displaying a clear cytoplasm and an accumulation of hyaline material testified to changes affecting their functional activities. Liver regeneration started properly, as early events initiating cell proliferation occurred within the first 3 h, and the G1 to S transition was detected in less than 12 h. However, a rise in p27 (Kip1) followed by p21 (Waf1/Cip1) cell cycle inhibitor levels led to a delayed S phase progression and mitosis. Overall, liver regeneration in rats with a 90% hepatectomy was delayed by 24 h and associated with a delayed onset and lower peak magnitude of hepatocellular deoxyribonucleic acid synthesis. CONCLUSIONThis work highlights the critical importance of the cyclin/cyclin-dependent kinase inhibitors of the Cip/Kip family in regulating the liver regeneration timeline following extended hepatectomy.     

Localization of Hsp27 in the Rat Submandibular Gland Following the Application of Various Surgical Treatments

Salivary glands repair and regenerate following various types of injuries and surgical procedures. However, the tissue responses induced in the contralateral glands have yet to be elucidated in detail. Hsp27, a member of the heat-shock protein (Hsp) family, is strongly expressed in physiological environments, particularly during development. Hsp27 was previously shown to play a role in the regulation of acinar cell proliferation and differentiation in the rat submandibular gland.The present study performed the following surgical treatments on the right submandibular glands of adult rats: 1) duct ligation followed by unligation after one week; 2) partial sialoadenectomy; and 3) total sialoadenectomy. Immunohistochemistry for Hsp27 and Ki67 was performed in the experimental and normal contralateral glands, and localization was histologically and morphometrically analyzed.The results obtained revealed the localization of Hsp27 to the intercalated duct in the submandibular glands of non-treated rats. The expression of Hsp27 was strongly induced in both the uninjured contralateral control glands as well as treated glands of experimental rats regardless of the surgical procedure performed. The number of Hsp27-immunopositive cells increased rapidly following surgery, and subsequently returned to the same level as that in non-treated rats after 4 weeks. However, no marked changes were observed in the number of Ki67-immunopositive proliferating cells. Therefore, the change in the number of Hsp27-immunopositive cells may have contributed to compensatory hypertrophy. The results of the present study indicate that the expression of Hsp27 in the intercalated duct in the submandibular gland may play a role in the differentiation of acinar cells.     

Evaluation of immunohistochemical expression of p53, p21, p27, cyclin D1, and Ki67 in oral and oropharyngeal squamous cell carcinoma

J Oral Pathol Med (2012) 41 : 40–46 Background: The purpose of this study was to evaluate whether the immunohistochemical expression of p53, p21, p27, cyclin D1, and Ki67 can predict therapy response and survival in patients with oral and oropharyngeal squamous cell carcinoma treated with preoperative chemoradiation. Methods: Biomarker expression was evaluated by immunohistochemistry in formalin‐fixed, paraffin‐embedded pretreatment biopsies of 111 homogenously treated patients. We assessed the association between clinicopathological variables including response to neoadjuvant chemoradiotherapy as well as the survival of the patients and the expression of the biomarkers as both dichotomized (positive vs. negative) and continuous variables. Results: Biomarker overexpression on the basis of pre‐selected cutoff points was seen in 66 of 111 (59%) cases for p53, in 77 (69%) for p21, in 48 (43%) for p27, in 81 (73%) for cyclin D1, and in 54 (49%) cases for Ki67, respectively. None of the examined biomarkers was able to predict response to neoadjuvant chemoradiotherapy or was associated with survival outcome. Post‐treatment pathologic TNM stage (P < 0.001), pathologic response (P < 0.001), and perineural invasion (P < 0.001) were the only factors having a significant effect on recurrence‐free survival. Post‐treatment pathologic N stage (P = 0.005), post‐treatment pathologic TNM stage (P < 0.001), pathologic response (P < 0.001), and perineural invasion (P = 0.001) had a significant impact on overall survival. Conclusions: Our results suggest that the biomarkers p53, p21, p27, cyclin D1, and Ki67 have no impact on treatment response and survival in patients with oral and oropharyngeal cancer treated with preoperative chemoradiation.     

Flow Cytometry Analysis with a New FITC-Conjugated Monoclonal Antibody-3E12 for HLA-B*57:01 Rapid Screening in Prevention of Abacavir Hypersensitivity in HIV-1–Infected Patients

Abstract

Background: Rapid screening for the detection of HLA-B*57:01 in the prevention of abacavir hypersensitivity in HIV-1–infected patients is a hallmark for clinical services. Objective: The aim of this work was to analyze the utility of flow cytometry with a new FITC-conjugated B-17 monoclonal antibody (mAb3E12) for HLA-B*57:01 screening in a Spanish cohort of 577 HIV-1+ individuals. Methods: Cryopreserved peripheral blood mononuclear cell samples from HIV-1+ individuals were analyzed by flow cytometry with the mAb 3E12 that recognizes both HLA-B*57 and HLA-B*58 alleles (members of the group specificity, HLA-B17). Patients’ DNA samples had been previously typed for HLA-B*57:01 with PCR-SSO or PCR-SSP and additional DNA sequencing (EPI Study). The results obtained by flow cytometry were compared with the results obtained by the DNA-PCR techniques. Results: By flow cytometry, 46 samples (7.97%) were positive for HLA-B17, 530 (91.86%) were negative, and 1 (0.17%) was undetermined. All samples found negative by flow cytometry were negative for HLA-B*57:01 by DNA-PCR. Of the HLA-B17 positive samples, 31 (67.4%) were positive for HLA-B*57:01, 2 (3.25%) were positive for HLA-B*57:03, 11 (26.1%) were positive for HLA-B*58, and 2 (3.25%) were negative for both HLA-B*57 and HLA–B*58 antigens. The undetermined sample was negative for HLA-B*57 and HLA-B*58 alleles by DNA-PCR. Conclusions: This study shows that flow cytometry with mAb3E12 is a highly sensitive method (no false negatives) to implement prior to DNA-PCR analysis for rapid screening of HLA-B*57:01. Additional confirmation by molecular HLA typing method would be required in less than 10% of the cohort of HIV-1–infected individuals.