Regulation of sperm function by reactive oxygen species

Sperm capacitation can be increased by the addition of reactive oxygen species (ROS) and decreased by antioxidants. Broadly consistent results have been achieved with a wide variety of methods and across different species. Exposure to ROS increases protein tyrosine phosphorylation consequent on an increase in cAMP and activation of tyrosine kinase and inhibition of tyrosine phosphatase. The measurement of ROS production by sperm is complicated by contamination of suspensions by leukocytes, laying many studies open to doubt. In human sperm the observation that extracellular NADPH could support superoxide production detected with the chemiluminescent probe lucigenin and had physiological effects similar to hydrogen peroxide led to the suggestion that they contained NADPH oxidase activity to generate ROS to support capacitation. However, the realization that lucigenin can signal superoxide artefactually, combined with failure to detect superoxide production using spin trapping techniques or to detect NADPH oxidase components in mature sperm, and confirmation of old reports that NADPH solution contains substantial amounts of hydrogen peroxide due to autoxidation, have undermined this hypothesis. Although the presence of significant NADPH oxidase activity in mature human sperm now seems less likely, other observations continue to suggest that they can make ROS in some way. There is stronger evidence that animal sperm can make ROS although these may be mainly of mitochondrial origin.     

Development of the NADPH-diaphorase-positive neurons in the sympathetic ganglia

Changes in the distribution of NADPH-diaphorase (NADPH-d) were studied in neurons of the superior cervical ganglion (SCG), stellate ganglion (SG) and celiac ganglia (CG) in newborn, 10-, 20-day-old, 1-month-old, 2-month-old and 6-month-old rats, mice and kittens. NADPH-d-positive neurons were revealed in all sympathetic ganglia in kittens but not in rodents from birth onwards. In kittens, the largest population of NADPH-d-positive cells was found in the SG, the smallest in the SCG (<1%) and we observed only a few cells in the CG. The proportion of NADPH-d-positive cells in the SG increased from 3.1±0.15% in newborn kittens to 9.3±0.63% in 20-day-old animals and decreased further from 8.1±0.75% in 30-day-old kittens to 3.4±0.54% in 2-month-old animals. The content of NADPH-d-positive cells in the CG and SCG did not change during development. There were no differences in cross-sectional area between neurons located in different ganglia of animals from the same age group under study. We conclude that the development of NADPH-d-positive neurons in different sympathetic ganglia has its own time dynamics and is completed by the end of the second month of life.     

Antimicrobial mechanisms of fish leukocytes

Early activation and coordination of innate defenses are critical for effective responses against infiltrating pathogens. Rapid engagement of immune cells provides a critical first line of defense soon after pathogen infiltration. Activation leads to a well-orchestrated set of events that sees the induction and regulation of intracellular and extracellular antimicrobial defenses. An array of regulatory mediators, highly toxic soluble molecules, degradative enzymes and antimicrobial peptides provides maximal protection against a wide range of pathogens while limiting endogenous damage to host tissues. In this review we highlight recent advances in our understanding of innate cellular antimicrobial responses of teleost fish and discuss their implications to cell survival, immunomodulation and death. The evolutionary conservation of these responses is a testament to their effectiveness against pathogen infiltration and their commitment to effective maintenance of host homeostasis. Importantly, recent developments in teleost fish systems have identified novel host defense strategies that may be unique to this lower vertebrate group or may point to previously unknown innate mechanisms that also play a significant role in higher vertebrate host immunity.     

运动干预糖尿病心肌病的分子机制研究

目的:观察链脲佐菌素诱导的糖尿病大鼠心肌组织中NADPH氧化酶表达的变化,探讨运动对NADPH氧化酶的影响。方法:大鼠糖尿病模型建立成功1周后,测定糖尿病对心肌组织中NADPH氧化酶亚基表达的影响,测定8周的运动能否有效影响NADPH氧化酶亚基的表达;观测糖尿病对心肌胶原蛋白表达的影响,8周运动是否能够影响胶原蛋白的表达。结果:糖尿病导致心室NADPH氧化酶亚基p47~(phox)和gp91~(phox)表达增加,而8周运动能够抑制这2种亚基表达的增加;糖尿病引起心房肌p47~(phox)表达显著增加,运动抑制其增加;糖尿病大鼠心肌胶原蛋白Ⅲ水平显著增加,运动降低胶原蛋白的增加。结论:运动减低糖尿病大鼠心脏p47~(phox)和gp91~(phox)的表达。这可能是改善心内基质的重要机制,因此运动干预糖尿病心肌病的一个有效机制可能是通过抑制心肌氧化应激和降低胶原蛋白的表达。     

How human neutrophils kill and degrade microbes: an integrated view

Summary:  Neutrophils constitute the dominant cell in the circulation that mediates the earliest innate immune human responses to infection. The morbidity and mortality from infection rise dramatically in patients with quantitative or qualitative neutrophil defects, providing clinical confirmation of the important role of normal neutrophils for human health. Neutrophil-dependent anti-microbial activity against ingested microbes represents the collaboration of multiple agents, including those prefabricated during granulocyte development in the bone marrow and those generated de novo following neutrophil activation. Furthermore, neutrophils cooperate with extracellular agents as well as other immune cells to optimally kill and degrade invading microbes. This brief review focuses attention on two examples of the integrated nature of neutrophil-mediated anti-microbial action within the phagosome. The importance and complexity of myeloperoxidase-mediated events illustrate a collaboration of anti-microbial responses that are endogenous to the neutrophil, whereas the synergy between the phagocyte NADPH (nicotinamide adenine dinucleotide phosphate) oxidase and plasma-derived group IIA phospholipase A₂ exemplifies the collective effects of the neutrophil with an exogenous factor to achieve degradation of ingested staphylococci.     

Nitroxide synthase activities at different stages of adjuvant arthritis

The location and activities of nitric oxide synthases in synovial cells during different stages of inflammatory process were studied by the immunocytochemical method in animals with experimental rheumatoid arthritis. Direct involvement of the NOergic mechanisms in the development of adjuvant arthritis was demonstarted. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 143, No. 5, pp. 576–579, May, 2007     

NADPH oxidase inhibition improves neurological outcomes in surgically-induced brain injury

Neurosurgical procedures can result in brain injury by various means including direct trauma, hemorrhage, retractor stretch, and electrocautery. This surgically-induced brain injury (SBI) can cause post-operative complications such as brain edema. By creating a mouse model of SBI, we tested whether NADPH oxidase, an important reactive oxygen species producing enzyme, is involved in SBI using transgenic mice lacking gp91^phox subunit of NADPH oxidase (gp91^phox KO) and apocynin, a specific inhibitor of NADPH oxidase. Neurological function and brain edema were evaluated at 24 h post-SBI in gp91^phox KO and wild-type littermates grouped into SBI and sham-surgery groups. Alternatively, mice were grouped into vehicle- and apocynin-treated (5 mg/kg, i.p. 30 min before SBI) groups. Oxidative stress indicated by lipid peroxidation (LPO) was measured at 3 and 24 h post-SBI. The gp91^phox KO mice, but not the apocynin-treated mice showed significantly improved neurological scores. Brain edema was observed in both gp91^phox KO and wild-type groups after SBI; however, there was no significant difference between these two groups. Brain edema was also not affected by apocynin-pretreatment. LPO levels were significantly higher in SBI group in both gp91^phox KO and wild-type groups as compared to sham group. A trend, although without statistical significance, was noted towards attenuation of LPO in the gp91^phox KO animals as compared to wild-type group. LPO levels were significantly attenuated at 3 h post-SBI by apocynin-pretreatment but not at 24 h post-SBI. These results suggest that chronic and acute inhibition of NADPH oxidase activity does not reduce brain edema after SBI. Long-term inhibition of NADPH oxidase, however improves neurological functions after SBI.     

Pentoxifylline modulates p47phox activation and downregulates neutrophil oxidative burst through PKA-dependent and -independent mechanisms

Background and Aim: Pentoxifylline (PTX) has been proven to be an inhibitor of fMLP-induced neutrophil (PMN) oxidative burst and is thought to function by increasing cAMP and Protein kinase A (PKA). We hypothesized that PTX diminishes production of the neutrophil respiratory burst by both PKA-dependent and independent mechanisms.

Material and Methods: Human neutrophils were isolated and stimulated with fMLP (1μM) alone or in combination with PTX (2mM). PMN activation was determined by the cytochrome C reduction method in the presence and absence of p38 MAPK (SB203580), ERK (PD98059), and PKA inhibitors (H89). Western blot analysis of Ras, Raf, p38 MAPK, ERK, and Akt was performed in PMNs exposed to fMLP and PTX. Cell membranes were fractionated to measure membrane-associated p47 phox. Treated cells were imaged using confocal microscopy to examine changes in localization of Akt and p47phox.

Results: PTX produced a decrease in oxidative burst that was diminished but not abrogated by H89 exposure. The reduction in Ras, Raf, and Akt activation seen with PTX was not effected by the presence of H89. The ability of PTX to attenuate phosphorylation of p38 MAPK and ERK was significantly decreased in the presence of H89, suggesting a PKA-dependent mechanisms. Membrane fractions of neutrophils demonstrate that PTX decreased membrane-associated p47phox, thus diminishing the ability to generate oxidative burst. PTX also decreased membrane localization of Akt and p47phox by confocal microscopy.

Conclusions: PTX attenuates activation of signaling molecules involved in activation of p47phox and suppress the subsequent assembly of the NADPH machinery through both PKA-dependent and PKA-independent mechanisms.     

NADPH氧化酶家族与高血压血管重构

高血压及其并发症的发生、发展与血管重构密切相关。高血压过程中血管重构的类型、特点以及发生机制已成为高血压研究领域的热点之一。高血压血管重构的机制尚未完全明了,研究表明活性氧诱导的氧化应激参与了高血压血管重构的各个环节。NADPH氧化酶是体内生成活性氧的主要酶类,大量研究证实NADPH氧化酶来源的ROS在高血压血管重构的发生、发展中起重要作用。本文就NADPH氧化酶介导高血压血管重构的作用及机制作一综述。     

雷公滕及铅、镉对小鼠睾丸间质细胞一氧化氮合酶活性的影响

目的　探讨雷公藤及铅、镉等重金属元素对睾丸间质细胞一氧化氮合酶 (NOS)活性的影响。　方法　用 NADPH- d黄递酶组织化学方法 ,观察给予氯化镉、醋酸铅、雷公藤 1、2、3、4周后 ,小鼠睾丸间质 NOS活性的变化。　结果　正常组间质细胞呈 NOS强阳性反应 ,而曲精小管生精细胞均呈 NOS阴性反应。给予雷公藤 2周内NOS反应无变化 ,第 3和第 4周后 ,NOS阳性的睾丸间质细胞数量无明显变化 ,但 NOS活性稍降低。给予醋酸铅和氯化镉 1周后 ,NOS阳性的睾丸间质细胞数量明显减少 ,并随给药时间延长而加重 ,NOS活性逐渐降低 ,给药 4周后 NOS阳性的睾丸间质细胞几乎不着色。　结论　雷公藤虽然具有明显抗生育作用 ,但对 NOS阳性的睾丸间质细胞数量及 NOS活性无明显影响 ;而铅、镉等对睾丸有毒理作用的重金属元素 ,不但可使 NOS阳性的睾丸间质细胞减少 ,NOS活性也明显降低。