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No description available.

G6Pase-alpha and G6Pase-beta share kinetic properties and active site structures, which lie on the luminal side of the endoplasmic reticulum (ER). For hydrolysis of G6P to glucose, G6Pase-alpha or G6Pase-beta must couple with an ubiquitously expressed ER-transmembrane protein, the G6P transporter (G6PT) that translocates G6P from the cytoplasm into the lumen of the ER. The primary role of the G6Pase/G6PT complex is therefore to provide endogenous glucose to the ER lumen. The essential role of the G6Pase-alpha/G6PT complex in glucose homeostasis has been well established, and the deficiencies in G6Pase-alpha and G6PT cause glycogen storage disease type Ia (GSD-Ia) and GSD-Ib, respectively. Both patients manifest the same metabolic phenotype of disturbed glucose homeostasis. While the metabolic abnormalities of GSD-Ia and GSD-Ib are almost identical, GSD-Ib patients exhibit neutropenia and myeloid dysfunctions which are not observed in GSD-Ia patients. Since G6Pase-beta and G6PT share an ubiquitous expression pattern, we hypothesized that the G6Pase-beta/G6PT complex might be functional in neutrophils and that the myeloid defects in GSD-Ib are due to the loss of activity of that complex. To test this hypothesis, we generated G6Pase-beta-deficient (G6pc3 --/--) mouse strains and showed that G6pc3--/-- mice manifest neutropenia; defects in neutrophil respiratory burst, chemotaxis, and calcium flux; and increased susceptibility to bacterial infection mimicking GSD-Ib patients. Consistent with this, G6pc3--/-- neutrophils exhibit enhanced ER stress and apoptosis. Taken together, the results demonstrate that endogenous glucose production in the ER via G6P translocation and metabolism are critical for normal neutrophil functions and that an ER stress-mediated neutrophil apoptosis is one mechanism underlying myeloid dysfunctions in the G6pc3--/-- mice. / Macrophages are the abundant leukocytes in the decidua throughout pregnancy and were thought to play a vital role in decidual homeostasis, placental development, and maintenance of a successful pregnancy. We hypothesized that endogenous glucose production in the ER might also be critical for normal macrophage function and G6pc3--/-- females manifesting neutropenia, neutrophil and macrophage dysfunctions might suffer from pregnancy-associated complications. Here we show that G6pc3--/-- macrophages exhibited impaired respiratory burst activity and repressed trafficking in vivo during an inflammatory response. The litter size and pregnancy frequency were markedly reduced in female G6pc3--/-- matings as compared to female G6pc3+/--/G6pc3+/+ matings, indicative of reduced fertility. The pregnancy-associated mortality risk was greatly increased in G6pc3--/--. Pathological analyses revealed that the sick or dying G6pc3--/-- mothers were emaciated and suffered from dental dysplasia and otitis media. Consistent with this, parental male and female G6pc3--/-- mice were more neutropenic than their age-matched virgin G6pc3 --/-- mice. Taken together, our results show that macrophage dysfunction, defective macrophage trafficking, neutrophil dysfunction, and enhanced neutropenia underlie the reduced fertility and increased mortality of G6pc3--/-- mothers. / Cheung, Yuk Yin. / Advisers: Janice Chou; Kam Bo Wong. / Source: Dissertation Abstracts International, Volume: 73-03, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 92-107). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

A total of 338 infants with suspected clinical sepsis were investigated, 115 of whom were found to be clinically infected. Twenty-one healthy term neonates were recruited as control subjects. The expression of CD64 on neutrophils in infected infants was significantly elevated at both 0 h and 24 h, compared with those of noninfected infants or controls (both p < 0.0005). The calculated optimal cutoff value for CD64 was 6136 antibody-phycoerythrin molecules bound/cell. CD64 has a very high sensitivity (96%) and NPV (97%) at 24 h. The use of CRP in combination with CD64 as predictive markers only marginally enhanced the sensitivity and NPV (97% and 98%, respectively). There was no statistical difference in the expression of monocyte HLA-DR among infected, noninfected, and control subjects. As a result, the optimal cutoff value for HLA-DR could not be determined. The technology of flow cytometry has potential applications for use in the diagnosis of neonatal sepsis because the measurement is quantitative, requiring only a minimal amount of whole blood and a short duration (within 3 h) for the provision of results. (Abstract shortened by UMI.) / Term newborns in whom infection was suspected when they were <72 h of age were recruited into the study. The expressions of CD64 on neutrophils and HLA-DR on monocytes were measured by flow cytometry at 0 h (the time of sepsis evaluation) and 24 h after the onset of presentation. A full sepsis screen, including complete blood count, serial C-reactive protein (CRP), blood culture, cerebrospinal fluid culture, and chest radiograph were performed. The demographic and clinical data were documented. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of neutrophil CD64, monocyte HLA-DR and the combination of markers for predicting neonatal sepsis were determined. / This prospective study aimed to evaluate the diagnostic utilities of two cell surface markers, neutrophil CD64 and monocyte HLA-DR, for the identification of early-onset clinical infection and pneumonia in term infants. The optimal cutoff value of each marker was defined according to the Receiver Operating Characteristic curve so that it could be used as a reference with which future studies can be compared. / Li Geng. / "May 2005." / Adviser: Pak Cheung Ng. / Source: Dissertation Abstracts International, Volume: 67-01, Section: B, page: 0174. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (p. 129-150). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Diss. (sammanfattning) Linköping : Univ., 2001. / Härtill 4 uppsatser.

Orientador: Sandra Helena Penha de Oliveira / Banca: Carlos Ferreira dos Santos / Banca: Pedro Felício Estrada Bernabé / Resumo: O objetivo deste trabalho foi investigar os mecanismos envolvidos na migração de neutrófilos induzida pelo MTA para cavidade peritoneal de camundongos. Observouse que o MTA induziu migração de neutrófilos de maneira dose-dependente (0,5; 5; 50 e 100 mg/cavidade), alcançando o pico de migração 6 horas após a injeção do estímulo com a dose de 50 mg/cavidade. Esta migração foi parcialmente inibida pelo pré-tratamento dos animais com dexametasona (1 mg/Kg), BWA4C (50 mg/Kg) e U75302 (0,5 mg/Kg). Diferentemente, a Indometacina (5 mg/Kg) foi inefetiva neste processo. Verificou-se também que os animais estimulados com MTA (50 mg/cavidade) apresentaram uma liberação significativa de IL-1ß e MIP-2 no exsudato peritoneal. O pré-tratamento com Tioglicolato aumentou em cerca de 380% a população de macrófagos na cavidade peritoneal, potencializando a migração de neutrófilos induzida pelo MTA (p<0,05). O pré-tratamento com composto 48/80 depletou cerca de 75% a população de mastócitos, diminuindo a migração de neutrófilos (p<0,05). A injeção de MTA na bolha de ar subcutânea induziu uma migração de neutrófilos menor comparada à cavidade peritoneal. Estes resultados confirmam a participação de mastócitos e macrófagos na migração de neutrófilos induzida pelo MTA. A injeção de sobrenadante de macrófagos e mastócitos estimulados com MTA na cavidade peritoneal de camundongos causou significante migração de neutrófilos (p<0,05), que foi parcialmente inibida pelo pré-tratamento das células por dexametasona (10 æMolar), BWA4C (100 æMolar) e U75302 (10 æMolar) sugerindo a liberação por essas células de LTB4 e citocinas e/ou quimiocinas. Confirmando esses dados,...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of this study was to investigate the mechanism involved in the neutrophil migration induced by MTA into peritoneal cavity in mice. It was observed that MTA induced a dose dependent neutrophil migration (0.5, 5, 50 and 100 mg/cavity), achieving the peak 6 hours after the stimulation with 50 mg/cavity. Neutrophil migration was inhibited by the pre-treatment with dexamethasone (1 mg/Kg), BWA4C (50 mg/Kg) and U75302 (0,5 mg/Kg). Differently indometacin (5 mg/Kg) was ineffective in this process. It was seen that the animals stimulated with MTA (50 mg/cavity) showed a significative amount of IL-1ß and MIP-2 released to the peritoneal exudate. The pretreatment with Thioglycolate 3% increased 380% the macropahge population into the peritoneal cavity, increasing the MTA-induced neutrophil migration (p<0.05). The pretreatment with 48/80 compound decreased 75% the mast cell population in the peritoneal cavity and decreased the MTAinduced neutrophil migration (p<0.05). The injection of MTA in the air-pouch cavity induced a neutrophil migration, however, the recruitment was shorter than that induced into the peritoneal cavity. These data confirm the participation of the mast cell and macrophages in the MTA-induced neutrophil migration. The injection of MTA-stimulated macrophages and mast cells supernatants into the mice peritoneal cavity induced a significant neutrophil migration that was inhibited by the pretreatment with dexamethasone (10 æMolar), BWA4C (100 æMolar) and U75302 (10 æMolar) suggesting the release of LTB4 and cytokines and/or chemokines by these cells. Besides, macrophages and mast cells MTA-induced were able to express in vitro IL-1ß MIP-2 and 5-LO mRNA. In conclusion, the neutrophil migration into mice peritoneal cavity induced by MTA was dependent on mast cells and macrophages, which expressed IL-1ß, MIP-2 and LTB4. / Mestre

Cathelicidins are cationic host defence peptides (CHDP) with essential roles in the innate defence system. These peptides have antimicrobial potential and the capacity to modulate innate immunity and inflammatory processes. Neutrophils (PMN) are the main reservoir of cathelicidins and play key roles in first line defence against infection. The appropriate regulation of PMN function, death, and clearance is critical to innate immunity, and the efferocytosis of apoptotic PMN, in contrast to necrotic cells, is proposed to promote the resolution of inflammation. In this thesis I demonstrate that the human cathelicidin LL-37 rapidly induced secondary necrosis of apoptotic human PMN and identify the essential C-terminal region of LL-37 required for this activity. In addition to the induction of secondary necrosis, higher concentrations of LL-37 also promoted PMN granule contents release. LL-37-induced secondary necrosis did not affect PMN ingestion by human monocyte-derived macrophages and, in contrast to expectation, was not proinflammatory. Interestingly, the anti-inflammatory effects of apoptotic PMN on activated macrophages were retained and even potentiated where LL-37-mediated secondary necrosis induced anti-inflammatory granule content release. Consistent with the results of in vitro studies, in vivo murine sterile peritonitis model revealed the same phenomenon: LL-37-induced secondary necrosis diminished inflammatory responses with decreased PMN influx. I also present data on LL-37- mediated modulation of innate immune effector cell cytokines responses to inflammatory signals. I propose that during acute inflammation LL-37 can modulate innate immune responses through its activity on cytokine production, and that LL-37-mediated secondary necrosis of apoptotic PMN has anti-inflammatory effects, but may also mediate host damage by promoting the release of potentially harmful intracellular contents under chronic or dysregulated conditions.

Background and Objectives: The pathophysiology of post-operative pneumonia following lung resection is poorly understood despite it being the most common complication which may lead to death. The role of the acute inflammatory response following lung resection, in particular innate immune cells, was investigated and used to identify biomarkers for post-operative pneumonia. Comparison of inflammatory responses to resection undertaken by video-assisted thorascopic surgery (VATS) and thoracotomy was also evaluated. Methods: Patients undergoing lung resection for suspected bronchogenic carcinoma were recruited. Objective pre-defined criteria were used to diagnose pneumonia. Bronchoalveolar lavage (BAL) was conducted in the contra-lateral lung pre- and postoperatively to measure cellular composition and cytokines. Blood was sampled preoperatively and 6-, 24- and 48-hours post-operatively primarily to assess neutrophil phagocytic capacity, monocyte subsets, monocyte cytokine responses to lipopolysaccharide (LPS) stimulation and serum cytokine responses. Exhaled nitric oxide (eNO) was also measured at these time points. Patient groups were compared using paired or student t-tests together with ANOVA/ANCOVA modelling. The predictive strength of the biomarkers identified was tested. Results: 40 patients were recruited. 26 patients (65%) underwent major lung resection using VATS and 14 (35%) thoracotomy. There was a post-operative blood monocytosis (p<0.0005) with an absolute expansion of classical and intermediate monocytes (p=0.001) and a relative fall in non-classical monocytes (p<0.005). Post-operatively blood monocytes became more pro-inflammatory with an overall significant increase in IL-8 (p=0.034) and TNF-α (p=0.028) together with an increase in IL-6 (p=0.028) and IL-10 by 48 hours (p=0.010). VATS was associated with a smaller release of IL-10 only (p=0.011). There was a general trend towards post-operative reduction in neutrophil phagocytosis of zymosan (in suspension) on ANOVA modelling (p=0.047). Lung resection led to an increase in serum cytokines IL-6, IL-8 and IL-10 which peaked at 24hrs before falling (p<0.0005). ANOVA modelling confirmed significantly lower levels of serum cytokines in VATS patients compared with thoracotomy (p=0.026 for IL-6, p=0.018 for IL-8 p=0.047 for IL-10). No significant post-operative change was found for IL-1β, TNF-α and IL-12p70 (p>0.05). Bronchoalveolar lavage fluid (BALF) and blood samples demonstrated a relative post-operative leucocytosis due principally to neutrophilia. A relative blood lymphopenia and thrombocytopenia developed postoperatively (p<0.0005). VATS was associated with a lower fall in serum albumin (p=0.001). BALF from the non-operated lung became more pro-inflammatory immediately post-operatively with an increase in IL-6 (p<0.0005), IL-8 (p=0.017), IL- 10 (p=0.018) and IL-1β (p=0.002). eNO tended to fall post-operatively which reached significance at 48 hrs (p=0.029). 14 patients developed pneumonia. Pre-operatively, a blood neutrophil count above 5.04x109/L had a relative risk (RR) for pneumonia of 3.3 (95% confidence interval (CI95) 1.1-10.1), and a BAL cell count of greater than 1.04x105/ml had a RR of 3.4 (CI95 1.3-9.0), whilst LPS-stimulated monocyte secretion of IL-12 of less than 0.15 pg/ml/μg protein had a RR of 3.0 (CI95 1.2-7.3). At 24 hours post-operatively, LPS-stimulated release from monocytes of IL-10 greater than 1.99 pg/ml/μg protein (RR 4.1, CI95 1.3- 12.3) and IL-6 greater than 414 pg/ml/μg protein (RR 3.1, CI95 1.2-8.1) were predictive of pneumonia. Conclusion: Lung resection is associated with significant early pro- and antiinflammatory responses. VATS resection invoked significantly lower levels of serum cytokines and albumin changes compared with thoracotomy suggesting VATS lobectomy should be the surgical treatment strategy of choice for early stage lung cancer. No difference in neutrophil function or monocyte function was however observed between the surgical groups. Clinical benefits of this reduced inflammation need to be evaluated in a larger cohort of patients. Relative pre-operative leucocytosis in blood and BAL together with monocyte hyper-responsiveness in the early postoperative period is associated with the development of pneumonia. These findings warrant further investigation for their predictive power in accurately identifying postoperative pneumonia. Ultimately, they may be incorporated into a risk stratification model enabling targeted prophylactic or earlier therapeutic intervention.

Neutrophils infiltrate myometrium and decidual tissue prior to parturition. Activated neutrophils release reactive oxygen species (ROS) and tumor necrosis factor α (TNFα), which might increase expression of pro-labor genes such as matrix metalloproteinase-1 (MMP-1), progesterone receptor (PR) A/B ratio, and cause demethylation of DNA. These changes might cause labor. Decidual tissue was obtained from consented, healthy women at term (37+ weeks of gestation) not in labor (no contractions, without cervical effacement), term labor and preterm labor (under 37 weeks of pregnancy). Decidual and myometrial cells in culture were treated with (1) ROS, (2) TNFα, or (3) 5-aza-2’-deoxycytidine. Total RNA was extracted, converted to cDNA and evaluated by qRT-PCR for MMP-1, PR-A+B and PR-B. TNFα increased MMP-1 by 17 fold in decidual cells and more than 12 fold in myometrial cells. PR-A/B was increased by 5.6 fold in decidua. ROS up-regulated MMP-1 by 6 fold and elevated the PR-A/B ratio by 4.5 fold in decidual tissue. DNA demethylation increased MMP-1 by about 4 and 11 fold in decidual and myometrium, respectively. The PR-A/B ratio was increased by 4 fold in decidua and the PR-B was decreased by 40% in the myometrium due to DNA demethylation. Decidual tissue in preterm labor showed a 7-fold increase in MMP-1 over term laboring and over a 15-fold increase over term not in labor tissue. In conclusion, MMP-1 expression and PR-A/B ratio was increased by neutrophil products possibly through a mechanism of DNA methylation in decidua and myometrium. Preterm decidua showed a dramatic increase in MMP-1 over normal labor tissue. TNFα and ROS increased expression of MMP-1 to possibly initiate parturition. These data might help explain mechanisms responsible for preterm labor unrelated to infection or premature rupture of membranes.