CCK-8对内毒素休克大鼠肺脏细胞因子的抑制效应

观察八肽胆囊收缩素(cholecystokinin-octapeptide,CCK-8）改善脂多糖(lipopolysaccharide,LPS）引起的大鼠内毒素性休克（endotoxic shock,ES）过程中血清及肺脏细胞因子的变化,探讨p38比裂素活化蛋白激酶（p38 mito-gen-activated protein kinase,p38 MAPK）的信号转导作用。用生理多道记录仪观察尾静脉注入LPS(p38 mito-gen-activated protein kinase,p38 MAPK）的信号转导作用。用生理多道记录仪观察尾静脉注入 LPS（8mg/kg i.v.）复制的SD大鼠ES模型、LPS注入前10min尾静脉注入CCK-8(40ug/kg i.v.）、单独注入CCK-8(40Uug/kg i.v.）或生理盐水（对照）的四组大鼠平均动脉血压（MAP）的改变,应用ELISA试剂盒检测血清和肺脏中炎性细胞因子（TNF-a、IL-1β和IL-6）的变化。用Western blot检测肺脏p38 MAPK的表达。结果显示：CCK-8可改善LPS引起的大鼠MAP的下降。与对照组相比,LPS可显著增加血清和肺脏TNF-a、IL-1β和IL-6含量;CCK-8可显著抑制LPS诱导的血清和肺脏TNF-a、IL-1β和IL-6的增加。CCK-8可增加ES大鼠肺脏磷酸化p38 MAPK的表达。结果提示CCK-8可改善ES大鼠MAP的降低,并对肺脏促炎性细胞因子过量产生有抑制作用,p38MAPK可能参与了其信号转导机制。     

肺内调节肽对兔支气管上皮细胞与嗜酸性粒细胞粘附功能的影响及机制探讨

为了探讨肺内调节肽在各类过敏性炎症发生,发展中的作用,我们观察了血管活性肠肽（vasoactive intestinal peptide,VIP)、表皮生长因子（epidermal growth factor,EGF)、内皮素-1（endothelin-1,ET-1)、降钙素基因相关肽（calcitonin gene-related peptide,CGRP)在未受应激与臭氧应激两种条件下对支气管上皮细胞（bronchial epithelial cell,BEC)与嗜酸性粒细胞（eosinophil,EOS)粘附的影响,结果发现,VIP、EGF可使O3应激的BEC与EOS的粘附率下降,下调气道上皮炎症反应：ET-1、CGRP可使未受应激的BEC与EOS的粘附率增加,诱发炎症损伤反应;CGRP还能加重臭氧的应激反应;ET-1、CGRP的效应可被W7、H7阻断,抗细胞间粘附分子-1（intercel-lular adhesion molecule,ICAM-1)抗体能阻断BEC与EOS的粘附,提示介导BEC与EOS粘附的粘附分子可能是ICAM-1。     

Human cytomegalovirus immediate early proteins and cell growth control

It is widely accepted that small DNA tumor viruses, such as adenovirus, simian virus 40 and papillomavirus, push infected cells into S-phase to facilitate the replication of their genome. Until recently, it was believed that the large DNA viruses (i.e. herpesviruses) functioned very differently in this regard by inducing a G₁ arrest in infected cells as part of their replication process. However, studies over the last 6–8 years have uncovered striking parallels (and differences) between the functions of the major immediate early (IE) proteins of at least one herpesvirus, human cytomegalovirus (HCMV) and IE equivalents encoded by small DNA tumor viruses, such as adenovirus. Similarities between the HCMV major IE proteins and adenovirus IE proteins include targeting of members of the RB and p53 families and an ability of these viral factors to induce S-phase in quiescent cells. However, unlike the small DNA tumor virus proteins, individual HCMV IE proteins target different RB family members. HCMV also encodes several other IE gene products as well as virion tegument proteins that act early during infection to prevent an infected cell from replicating its host genome and from undergoing apoptosis. Here, we review the specifics of several HCMV IE proteins, two virion components, and their functions in relation to cell growth control.     

Effects of cell swelling on intracellular calcium and membrane currents in bovine articular chondrocytes

Chondrocytes experience a dynamic extracellular osmotic environment during normal joint loading when fluid is forced from the matrix, increasing the local proteoglycan concentration and therefore the ionic strength and osmolarity. To exist in such a challenging environment, chondrocytes must possess mechanisms by which cell volume can be regulated. In this study, we investigated the ability of bovine articular chondrocytes (BAC) to regulate cell volume during a hypo-osmotic challenge. We also examined the effect of hypo-osmotic stress on early signaling events including [Ca2+](i) and membrane currents. Changes in cell volume were measured by monitoring the fluorescence of calcein-loaded cells. [Ca2+](i) was quantified using fura-2, and membrane currents were recorded using patch clamp. BAC exhibited regulated volume decrease (RVD) when exposed to hypo-osmotic saline which was inhibited by Gd3+. Swelling stimulated [Ca2+](i) transients in BAC which were dependent on swelling magnitude. Gd3+, zero [Ca2+](o), and thapsigargin all attenuated the [Ca2+](i) response, suggesting roles for Ca2+ influx through stretch activated channels, and Ca2+ release from intracellular stores. Inward and outward membrane currents significantly increased during cell swelling and were inhibited by Gd3+. These results indicate that RVD in BAC may involve [Ca2+](i) and ion channel activation, both of which play pivotal roles in RVD in other cell types. These signaling pathways are also similar to those activated in chondrocytes subjected to other biophysical signals. It is possible, then, that these signaling events may also be involved in a mechanism by which mechanical loads are transduced into appropriate cellular responses by chondrocytes.     

Expression of a releasable form of annexin II by human keratinocytes

Annexin II is a multifunctional calcium-dependent phospholipid binding protein whose presence in epidermis has previously been reported. However, like other members of annexin family, annexin II has been regarded as either an intracellular protein or associated with the cellular membrane. Here, we report the presence of a releasable annexin II and p11, two monomers of annexin II tetramer, in keratinocyte-conditioned medium (KCM). Proteins present in KCM were fractionated on a gel filtration column and following further evaluation, a releasable protein with apparent MW of 36 kDa was identified. Further characterization identified this protein as the p36 monomer of annexin II tetramer. The phospho-tyrosine antibody did not visualize this protein as the phosphorylated form of p36. Several experiments were conducted to examine whether this protein is soluble or associated with keratinocyte cell membranes in the conditioned medium. A centrifugation of conditioned medium was not able to bring this protein down into the pellet. Surprisingly, the results of Western analysis identified p36 and p11, two monomers of the annexin II tetramer, in conditioned medium derived from either keratinocytes cultured alone or keratinocytes co-cultured with fibroblasts. In contrast to the keratinocyte-conditioned medium in which annexin II was easily detectable, both monomers were barely detectable in conditioned medium collected from dermal fibroblasts. This finding was in contrast to the cell lysates in which p36 was detectable in both keratinocytes and fibroblasts. However, the amount of this protein was markedly higher in keratinocyte lysate relative to that of dermal fibroblasts. Conditioned medium derived from keratinocyte established from adult showed a higher level of annexin II compared to that of keratinocytes established from newborn babies. The expression of p11 seems to increase with differentiation of keratinocytes derived from either adult or newborn skin samples. When the site of annexin synthesis in human skin was examined by immunohistochemical staining, the antibody for p36 localized the annexin to the keratinocyte cell members in the basal and suprabasal keratinocytes. In conclusion, Western blot detection of both p36 and p11 in conditioned medium from skin cells revealed that human keratinocytes, but not fibroblasts, express a releasable monomer form of annexin II which is regulated by differentiation status of keratinocytes. This finding is consistent with the localization of annexin II detected by immunohistochemical staining.     

Cadmium-induced apoptosis of primary epithelial lung cells: Involvement of Bax and p53, but not of oxidative stress

The lung is a target organ for cadmium (Cd) toxicity. Apoptosis induced by cadmium acetate (CdAc) was studied in alveolar type 2 cells and Clara cells isolated from rat lung. Relatively low concentrations of CdAc (1–10 μmol/L) induced apoptosis after exposure for 20 h. Type 2 cells were more sensitive than Clara cells to Cd-induced apoptosis and loss of cell viability. On exposure to 10 μmol/L CdAc, the levels of the apoptosis-modulating proteins p53 and Bax were increased at 2 h and 5–12 h, respectively. The expression of p53 preceded the expression of Bax and the apoptotic process. The exposure to 10 μmol/L CdAc did not significantly increase the formation of cellular reactive oxygen species (ROS). However, after exposure to a high concentration of CdAc (100 μmol/L), a 30% increase of the ROS level was observed. No significant nitric oxide production was measured following CdAc exposure. Catalase, superoxide dismutase, dimethyl sulfoxide, or tetramethylthiourea did not protect against Cd-induced apoptosis. In conclusion, the results show that Clara cells and type 2 cells are sensitive to Cd-induced apoptosis. Increased levels of p53 and Bax are suggested to be involved in the apoptosis. The apoptosis did not appear to be mediated by oxidative pathways. This revised version was published online in August 2006 with corrections to the Cover Date.     

Involvement of p38 MAP kinase during iron chelator-mediated apoptotic cell death

Iron is an essential element for the neoplastic cell growth, and iron chelators have been tested for their potential anti-proliferative and cytotoxic effects. To determine the mechanism of cell death induced by iron chelators, we explored the pathways of the three structurally related mitogen-activated protein (MAP) kinase subfamilies during apoptosis induced by iron chelators. We report that the chelator deferoxamine (DFO) strongly activates both p38 MAP kinase and extracellular signal-regulated kinase (ERK) at an early stage of incubation, but slightly activates c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK) at a late stage of incubation. Among three MAP kinase blockers used, however, the selective p38 MAP kinase inhibitor SB203580 could only protect HL-60 cells from chelator-induced cell death, indicating that p38 MAP kinase serves as a major mediator of apoptosis induced by iron chelator. DFO also caused release of cytochrome c from mitochondria and induced activation of caspase 3 and caspase 8. Interestingly, treatment of HL-60 cells with SB203580 greatly abolished cytochrome c release, and activation of caspase 3 and caspase 8. Collectively, the current study reveals that p38 MAP kinase plays an important role in iron chelator-mediated cell death of HL-60 cells by activating downstream apoptotic cascade that executes cell death pathway.     

Role of cell cycle regulator p19ARF in regulating T cell responses

Although it is well established that the processes of cellular proliferation and apoptosis are linked, the role of cell cycle regulators in T cell responses in vivo is not well understood. In recent years, tumor suppressor molecule p19(ARF) has emerged as a key cell cycle regulator important in cellular apoptosis against strong mitogenic stimuli. In this study, we compared the antigen-specific T cell responses between wild type (+/+) and p19(ARF)-deficient (p19-/-) mice following an acute infection with lymphocytic choriomeningitis virus (LCMV). p19-/- mice mounted a potent CD8 T cell response and the magnitude of expansion of LCMV-specific CD8 T cells was comparable to that of +/+ mice. Further, the clonal downsizing of the expanded virus-specific CD8 T cells and establishment of long-term T cell memory were minimally affected by p19(ARF) deficiency. Therefore, p19(ARF) function is not essential to regulate T cell responses following an acute viral infection.