Age-related dysfunction of p53-regulated phagocytic activity in macrophages.

Aging promotes polarization of M2-like macrophages to M1-like macrophages and reduces their phagocytic ability. However, the molecular mechanisms underlying these aging-related changes remain poorly understood. Here, we demonstrate that p53 regulates phagocytic activity in macrophages from young mice but not in those from old ones. Macrophages from both old and young mice expressed functional p53 to induce target genes including p21 and Mdm2. In macrophages from young mice, chemically induced p53 decreased phagocytic activity and c-Myc levels, with the latter change reducing M2-related genes. However, in macrophages from old mice, phagocytic activity and c-Myc expression were independent of p53 activity. Furthermore, c-Myc suppression did not affect M2-related genes in old-mouse macrophages. These results demonstrate that dysregulation of p53 function is a molecular mechanism underlying reduced phagocytic activity in aged-mouse macrophages.

Current Trends of Neutrophil Biology.

Neutrophils are short-lived and terminally differentiated cells, and therefore, have been considered as effector cells to phagocytose pathogens and kill them or damage tissues [...].

Kinetics of cytokine mRNA and protein expression by plastic adherent cells in the thymus after split-dose irradiation.

Whole body irradiation causes significant apoptosis in various tissues such as the thymus. If apoptotic cells outnumber the phagocytic capacity of macrophages, apoptosis becomes secondary necrosis, inducing inflammatory cytokine expression in macrophages. Radiation also induces thymic lymphomas in C57BL/6 mice after four consecutive irradiations with 1.6 Gy X-rays with nearly 100% incidence. Since cancer development is modulated by a microenvironment involving macrophages, we examined the kinetics of thymocyte number and plastic adherent cell number in the thymus as well as cytokine mRNA expression by plastic adherent cells in the thymus after split-dose irradiation. Upon split-dose irradiation, thymocyte number changed dramatically, whereas plastic adherent cell number did not. Among cytokine mRNAs tested, IL-1ß, IL-11 and IL-12p40 mRNAs were up regulated 2 days after the 1st and 2nd, 3rd and 4th, and 2nd and 3rd irradiations, respectively. On the other hand, TNF-α mRNA was up regulated 2 days after the 3rd irradiation and 2 weeks after the 4th irradiation. The level of IL-11 protein was also increased 2 days after 3rd and 4th irradiations. These results suggest that, upon split-dose irradiation, macrophages in the thymus produce various cytokines in a time-dependent manner, thereby contributing to induction of thymic lymphomas.

Impact of Left Heart Bypass on Arterial Oxygenation During One-Lung Ventilation for Thoracic Aortic Surgery.

OBJECTIVE: The aim of this study was to reveal the mechanism of improved arterial oxygenation by measuring the changes in oxygenation before and after initiation of left heart bypass (LHB) during one-lung ventilation (OLV) for thoracic aortic surgery. DESIGN: Prospective, observational study. SETTING: Single-institution, private hospital. PARTICIPANTS: The study comprised 50 patients who underwent aortic surgery via a left thoracotomy approach with LHB circulatory support. INTERVENTIONS: Patients were ventilated using pure oxygen during OLV, and the ventilator setting was left unchanged during the measurement period. MEASUREMENTS AND MAIN RESULTS: The measurement of partial pressure of arterial oxygen (PaO2) was made at the following 4 time points: 2 minutes after heparin infusion (point 1 [P1]), 2 minutes after inflow cannula insertion through the left pulmonary vein (P2), immediately before LHB initiation (P3), and 10 minutes after LHB initiation (P4). The mean±standard deviation (mmHg) of PaO2 measurements at the P1, P2, P3, and P4 time points were 244±121, 250±123, 419±122, and 430±109, respectively, with significant increases between P1 and P3, P1 and P4, P2 and P3, and P2 and P4 (p<0.0001, respectively). No significant increase in PaO2 was seen between P1 and P2 or between P3 and P4. CONCLUSIONS: The improved arterial oxygenation during OLV in patients who underwent thoracic aortic surgery using LHB can be attributed to the insertion of an inflow cannula via the left pulmonary vein into the left atrium before LHB.

Skewing of peritoneal resident macrophages toward M1-like is involved in enhancement of inflammatory responses induced by secondary necrotic neutrophils in aged mice.

Secondary necrotic cells, which are generated if apoptotic cells are incompletely cleared, induce severe inflammatory responses involving MIP-2 production and subsequent neutrophil infiltration. Recently, we showed that the phagocytic capacity of peritoneal resident macrophages from wild type (WT) aged mice as well as SMP30(-/-) mice fed a VC-limited diet as to secondary necrotic cells was reduced as compared with that in young mice, and that the inflammatory responses induced were stronger than those in young mice, presumably because of the delay in removal of secondary necrotic cells in aged mice. In this study, we investigated why MIP-2 production was increased in aged mice upon injection of secondary necrotic cells and why the phagocytic capacity of peritoneal resident macrophages from aged mice was reduced. When cocultured with secondary necrotic cells, the peritoneal resident macrophages from both types of aged mice significantly produced MIP-2 even in the absence of IFN-γ, whereas MIP-2 production by macrophages from WT young mice required IFN-γ. The peritoneal resident macrophages from both types of aged mice expressed CD40, a M1 macrophage marker, as in the case of M1 macrophages, which were obtained by treatment of macrophages from WT young mice with IFN-γ and LPS. Furthermore, M1 macrophages exhibited less phagocytic capacity as to secondary necrotic cells than non-treated macrophages. These results suggest that the phenotype of peritoneal resident macrophages is skewed toward M1-like in aged mice and that such skewing toward M1-like is involved in enhancement of inflammatory responses induced by secondary necrotic neutrophils in aged mice.

[Hyperthyroidism Diagnosed from Refractory Tachycardia and Hypotension during Surgery: A Case Report].

We report a case of hyperthyroidism diagnosed from refractory tachycardia and hypotension during surgery. Although the patient had exhibited tachycardia preop- eratively, it was difficult to suspect hyperthyroidism due to specific conditions of neurosurgical patient Eventually diagnosis of hyperthyroidism was made by exclusion, and prompt treatment was effectively initiated. Recently several reports suggested that landiolol was effective for rate control in patients with hyperthyroidism. At first intravenous bolus doses of landiolol were administered but were insufficient Secondly, intravenous propranolol was administered and tachycardia as well as blood pressure improved. The benefits of propranolol has been suggested.

Integrin-αvß3 regulates thrombopoietin-mediated maintenance of hematopoietic stem cells.

Throughout life, one's blood supply depends on sustained division of hematopoietic stem cells (HSCs) for self-renewal and differentiation. Within the bone marrow microenvironment, an adhesion-dependent or -independent niche system regulates HSC function. Here we show that a novel adhesion-dependent mechanism via integrin-ß3 signaling contributes to HSC maintenance. Specific ligation of ß3-integrin on HSCs using an antibody or extracellular matrix protein prevented loss of long-term repopulating (LTR) activity during ex vivo culture. The actions required activation of αvß3-integrin "inside-out" signaling, which is dependent on thrombopoietin (TPO), an essential cytokine for activation of dormant HSCs. Subsequent "outside-in" signaling via phosphorylation of Tyr747 in the ß3-subunit cytoplasmic domain was indispensable for TPO-dependent, but not stem cell factor-dependent, LTR activity in HSCs in vivo. This was accompanied with enhanced expression of Vps72, Mll1, and Runx1, 3 factors known to be critical for maintaining HSC activity. Thus, our findings demonstrate a mechanistic link between ß3-integrin and TPO in HSCs, which may contribute to maintenance of LTR activity in vivo as well as during ex vivo culture.

Nonocclusive mesenteric ischemia following multiple wasp stings.

Massive wasp envenomation can cause not only severe immediate allergic reactions and anaphylaxis but also severe delayed toxin-mediated systemic reactions, including hemolysis, coagulopathy, rhabdomyolysis, acute renal failure, and hepatotoxicity. However, reports of the latter type of reactions are rare. The subject of this case report, a 66-year-old man, was stung more than 30 times during an attack by wasps. Although he initially complained of pain, he showed no signs of anaphylaxis during observation in an emergency department. Twenty hours after envenomation, he was admitted to the hospital because of vomiting, abdominal pain, and lower gastrointestinal bleeding. Mesenteric ischemia, rhabdomyolysis, acute renal failure, and hepatotoxicity were diagnosed as delayed toxin-mediated systemic reactions resulting from massive wasp envenomation. Contrast-enhanced computed tomography findings, which included no thrombi or emboli but did reveal the abrupt tapering of mesenteric arteries, strongly suggested that the ischemia was due to nonocclusive mesenteric ischemia. Immediately after diagnosis, an emergency laparotomy was performed. Nonocclusive mesenteric ischemia was finally diagnosed via a histologic examination of the resected small bowel. We present the first case report of nonocclusive mesenteric ischemia consequent to wasp stings.

[Analgesia with paravertebral block for postoperative pain after thoracic or thoracoabdominal aortic aneurysm repair].

Paraplegia is a serious complication after thoracoabdominal aortic aneurysm repair. Therefore, maintenance of spinal cord perfusion pressure, drainage of cerebrospinal fluid, and avoidance of opioids are important for prevention of paraplegia Management of acute post-thoracotomy pain is necessary not only to keep the patient comfortable but also to minimize postoperative complications. However, epidural analgesia, a common method of pain control, is hard to use because of existing postoperative coagulopathy and avoidance of spinal cord ischemia Although both paravertebral block and epidural analgesia provide comparable pain relief after thoracic surgery, paravertebral block has lesser detrimental effects on spinal cord perfusion and better preserves the possibility to monitor neurologic function than epidural analgesia. We report 7 cases in which paravertebral blockade was used for analgesia in patients who underwent thoracoabdominal aneurysm repair.

Suppression of MIP-2 or IL-8 production by annexins A1 and A4 during coculturing of macrophages with late apoptotic human peripheral blood neutrophils.

Annexin A1 (ANXA1) is a well-known anti-inflammatory protein that is expressed on the surface of apoptotic cells. Annexin A4 (ANXA4) is also recruited from the nucleus to the cytoplasm in apoptotic cells, although it is not known whether or not ANXA4 is expressed on the surface of apoptotic cells. In this study, we obtained rabbit anti-human ANXA1 and ANXA4 antibodies, and then examined whether or not ANXA1 and ANXA4 are expressed on the surface of early and late human apoptotic cells. ANXA1 and, to a lesser extent, ANXA4 were detected on late but not early apoptotic HeLa cells, whereas ANXA1 and a small amount of ANXA4 were detected on both early and late apoptotic human neutrophils. We then examined the effects of the anti-human ANXA1 and ANXA4 antibodies on the mouse or human macrophage response to human apoptotic cells. Upon coculturing of mouse or human macrophages with late apoptotic human neutrophils, anti-human ANXA1 antibodies and, to a lesser extent, anti-human ANXA4 antibodies increased MIP-2 or IL-8 production significantly, suggesting that ANXA1 and ANXA4 suppress MIP-2 or IL-8 production by macrophages in response to late apoptotic human neutrophils.

Effects of regular exercise on neutrophil functions, oxidative stress parameters and antibody responses against 4-hydroxy-2-nonenal adducts in middle aged humans.

Regular exercise has recognized health benefits, partly because it reportedly lowers the levels of the oxidation products of proteins and DNA at rest, in contrast with the effect of acute exercise. However, when we compared oxidative response markers in active middle-aged subjects with those in sedentary ones, the level of urinary 8-OHdG was higher in active subjects. Because neutrophils are the first line of defense against a variety of infectious diseases, we then compared the cell density, functions and apoptosis of neutrophils in active subjects with those in sedentary ones. The cell density of neutrophils and phagocytosis of opsonized zymosan by neutrophils were higher in active subjects, being similar with the reported effects of acute exercise. To determine any beneficial effects of oxidative stress in active subjects, we then compared the levels of antibodies against 4-hydroxy-2-nonenal adducts in active subjects with those in sedentary ones, because 4-hydroxy-2-nonenal is one of the most common bioactive aldehyde products of oxidative stress, and because the IgM class of antibodies against oxidized low-density lipoprotein is associated with atheroprotective properties. The level of the IgM but not the IgG class of antibodies against 4-hydroxy-2-nonenal adducts was higher in active subjects. Overall, this study revealed that our active middle-aged subjects showed both oxidative responses and a higher IgM response to reactive carbonyl derivatives, possibly providing a basis for a health benefit by exercise in our active subjects.

Regulation of the estrous cycle by neutrophils via opioid peptides.

We found previously that neutrophil-depleted mice exhibited significant blockading of both the regular estrous cycle and cyclic changes of steroid hormone levels. In this study, we aimed at elucidation of the underlying mechanism. To examine the possibility that an increase in bacteria in the vaginal vault of neutrophil-depleted mice causes blockading of the estrous cycle, we treated neutrophil-depleted mice with antibiotics but failed to restore the estrous cycle. We then examined another possibility that neutrophils regulate the estrous cycle via opioid peptides, because opioid peptides regulate steroidogenesis in theca and granulosa cells in the ovaries, and because neutrophils contain opioid peptides. In support of this possibility, naloxone, an opioid antagonist, blocked the estrous cycle and a µ opioid receptor agonist restored the estrous cycle in neutrophil-depleted mice. Pro-opiomelanocortin was immunohistochemically detected in peripheral blood neutrophils but not in ones that had infiltrated into the ovaries. i.v. injection of anti-MIP-2 polyclonal Ab caused blockading of the estrous cycle, whereas MIP-2 was detected in the ovaries, suggesting a role of MIP-2 in the regulation of the estrous cycle. Moreover, i.v. injection of MIP-2 decreased the pro-opiomelanocortin signal in peripheral blood neutrophils and caused blockading of the estrous cycle. Together, these results suggest that neutrophils maintain the estrous cycle via opioid peptides.

Interleukin-9 receptor gene is transcriptionally regulated by nucleolin in T-cell lymphoma cells.

Interleukin-9 (IL-9) is a multifunctional cytokine that not only has roles in immune and inflammatory responses but also is involved in growth-promoting and anti-apoptotic activities in multiple transformed cell lines, which suggests a potential role in tumorigenesis. Over-expression of the receptor of IL-9 (IL-9R) occurs in several types of human leukemias and in radiation-induced mouse T-cell lymphoma (TL). The molecular mechanism that regulates transcription of the IL-9R gene (Il9r) during leukemogenesis is, however, not well understood. Using a mouse TL cell line that has high expression of Il9r, we sought to dissect its promoter structure. Here we show that the active promoter for Il9r is located in the 5'-flanking AT-rich region. Chromatin immunoprecipitation showed the opening of chromatin structure of the promoter region coupled with nucleolin binding in vivo. Immunohistochemical analysis confirmed the increased localization of nucleolin in the nuclei of TL cells. These data indicate that increased expression of Il9r is associated with an increased binding of nucleolin, coupled with chromatin opening, to an AT-rich region in the 5'-flanking region of Il9r in TL cells.

The mechanism underlying the appearance of late apoptotic neutrophils and subsequent TNF-α production at a late stage during Staphylococcus aureus bioparticle-induced peritoneal inflammation in inducible NO synthase-deficient mice.

During inflammation, neutrophils infiltrate into the involved site and undergo apoptosis. Early apoptotic neutrophils are then cleared by phagocytes, leading to resolution of the inflammation, whereas if late apoptotic neutrophils are accumulated for some reason, they provoke proinflammatory responses such as TNF-α production. To determine how endogenously produced nitric oxide (NO) regulates neutrophil apoptosis and the resolution of inflammation, we compared peritoneal inflammation induced by Staphylococcus aureus bioparticles in wild type mice with that in inducible NO synthase (iNOS)-deficient ones. In this model, NO production was largely dependent on iNOS, the NO level peaking at 24 h. There were increases in the numbers of neutrophils and late apoptotic ones at 24 h in iNOS-deficient mice as compared with in wild type ones, and consequently TNF-α production at 36 h in iNOS-deficient mice. On the other hand, the administration of a NO donor to iNOS-deficient mice at 12 h decreased the numbers of neutrophils and late apoptotic ones at 24 h, and thereafter TNF-α production at 36 h. In addition, coculturing of macrophages with late apoptotic neutrophils caused TNF-α production and a NO donor inhibited the transmigration of neutrophils in a dose-dependent manner. Collectively, these results suggest a novel mechanism that endogenously produced NO suppresses neutrophil accumulation at a late stage of inflammation, thereby preventing the appearance of late apoptotic neutrophils and subsequent proinflammatory responses.

The importance of infiltrating neutrophils in SDF-1 production leading to regeneration of the thymus after whole-body X-irradiation.

Our previous study demonstrated that neutrophils transiently infiltrated into a site where apoptosis had been induced. However, the role of infiltrating neutrophils has not been fully elucidated. In this study, we examined their role in regeneration of the thymus after whole-body X-irradiation by focusing on SDF-1 production. After X-irradiation, the thymus became severely atrophied presumably due to phagocytosis of apoptotic thymocytes. At that time, a significant number of neutrophils were detected in the thymus. The thymus was then partially regenerated on day 7, whereas the level of SDF-1 in it was significantly increased on days 3 and 5. Depletion of neutrophils greatly impaired SDF-1 production and the thymus regeneration. Moreover, administration of a CXCR4 antagonist also greatly suppressed the thymus regeneration. Furthermore, coculturing of a stromal cell line with infiltrating neutrophils increased SDF-1 production. These results suggest that infiltrating neutrophils play an auxiliary role in regeneration of the thymus after whole-body X-irradiation through augmentation of SDF-1 production.

Apoptotic neutrophils and nitric oxide regulate cytokine production by IFN-γ-stimulated macrophages.

Early apoptotic neutrophils but not secondary necrotic ones down-regulate LPS-induced proinflammatory cytokine production of macrophages, thereby contributing to the resolution of inflammation. IFN-γ is also a well-known stimulant of macrophages, but how the apoptotic neutrophils affect IFN-γ-stimulated macrophages remains largely unexplored. Since IFN-γ induces the expression of inducible nitric oxide (NO) synthase, we examined the production of NO and various cytokines, including MIP-2, TNF-α, IL-12p40, IL-6, IL-10, and TGF-ß, by IFN-γ-stimulated murine macrophages, the effect of coculturing the macrophages with early apoptotic or secondary necrotic neutrophils, and the regulatory role of NO in such cocultures. IFN-γ induced significant production of NO, IL-12p40, and IL-6 by macrophages, but not other cytokines. Early apoptotic neutrophils but not secondary necrotic ones promoted NO production, whereas secondary necrotic ones and their supernatants promoted TNF-α production. In contrast, both early apoptotic and secondary necrotic neutrophils suppressed IL-12p40 and IL-6 production. Furthermore, macrophages from inducible NO synthase-deficient mice produced significantly higher levels of MIP-2 than those from wild-type mice. Consistent with this, treatment of macrophages with l-NAME, an NO synthase inhibitor, also induced the production of a large amount of MIP-2. In conclusion, this study suggests that early apoptotic neutrophils are critical in the resolution of inflammation, but that secondary necrotic neutrophils may not cause an inflammatory response. Apoptotic neutrophils, however, appear not to modulate cytokine production via NO.

Mechanism underlying silent cleanup of apoptotic cells.

Apoptotic cells are cleared without an inflammatory response such as neutrophil infiltration. The mechanism underlying such silent cleanup of apoptotic cells has been intensively investigated in vitro for over a decade, and the concept that active suppression via IL-10, TGF-ß, and nitric oxide enables such silent cleanup to occur has been emerging. However, because this concept has not been vigorously examined under a variety of experimental conditions in vivo, the possibility remains that a null response, in which neither cytokines nor nitric oxide is produced upon an encounter with apoptotic cells, is responsible for silent cleanup.

Acute kidney injury during aortic arch surgery under deep hypothermic circulatory arrest.

PURPOSE: The aim of this investigation was to describe the renal outcome and to identify risk factors for acute kidney injury (AKI), as defined by the Acute Kidney Injury Network (AKIN), during aortic arch surgery (AAS) under deep hypothermic circulatory arrest (DHCA). METHODS: A retrospective and observational study has been performed. One hundred thirty-five patients requiring AAS under DHCA were studied. RESULTS: Seventy-one patients (52.6%) developed AKI during the postoperative period. A logistic regression analysis identified three independent risk factors for AKI: preoperative hypertension (HT), emergency surgery, and duration of DHCA. Renal replacement therapy (RRT) was required in four patients (3.0%). The postoperative mortality rate among the patients with AKI was 2.8%, which was not statistically different from the rate of 1.6% observed in the non-AKI group (P = 0.62). CONCLUSIONS: A high incidence of AKI during AAS under DHCA was confirmed. Because AKI is highly associated with aortic surgery, novel approaches for protecting the kidneys other than deep hypothermia are needed. The logistic regression model identified HT, emergency surgery, and duration of DHCA as independent risk factors for AKI.

Complicated biallelic inactivation of Pten in radiation-induced mouse thymic lymphomas.

Inactivation of the phosphatase and tensin homolog gene (Pten) occurs via multiple tissue-dependent mechanisms including epigenetic silencing, point mutations, insertions, and deletions. Although frequent loss of heterozygosity around the Pten locus and plausible involvement of epigenetic silencing have been reported in radiation-induced thymic lymphomas, the proportion of lymphomas with inactivated Pten and the spectrum of causal aberrations have not been extensively characterized. Here, we assessed the mode of Pten inactivation by comprehensive analysis of the expression and alteration of Pten in 23 radiation-induced thymic lymphomas developed in B6C3F1 mice. We found no evidence for methylation-associated silencing of Pten; rather, complex structural abnormalities comprised of missense and nonsense mutations, 1- and 3-bp insertions, and focal deletions were identified in 8 of 23 lymphomas (35%). Sequencing of deletion breakpoints suggested that aberrant V(D)J recombination and microhomology-mediated rearrangement were responsible for the focal deletions. Seven of the 8 lymphomas had biallelic alterations, and 4 of them did not express Pten protein. These Pten aberrations coincided with downstream Akt phosphorylation. In conclusion, we demonstrate that Pten inactivation is frequently biallelic and is caused by a variety of structural abnormalities (rather than by epigenetic silencing) and is involved in radiation-induced lymphomagenesis.

[Anesthetic management of a patient with chronic spinal cord injury for laparoscopic cholecystectomy].

Spinal cord injury (SCI) is a rare disease that offers challenges to anesthesiologists, while laparoscopic cholecystectomy (LC) has become common in recent years. We report a case of adult patient with chronic high SCI who underwent LC. A 62-year-old man, a known case of cervical SCI, was presented for LC. Anesthetic problems included circulatory and respiratory complications because of both SCI and pneumoperitoneum. Anesthesia was induced with propofol and a standard endotracheal tube was inserted with vecuronium; and thereafter anesthesia was maintained with small bolus doses of fentanyl and sevoflurane inhalation in the absence of epidural block. The intra- and post-operative course was completely uneventful without any episode of autonomic hyperreflexia. Due to a lack of sensory and motor function, SCI patients will receive little benefit from minimally invasive laparoscopic procedures. In conclusion, compared to open laparotomy, LC will minimize surgical trauma and hospital stay, but may not always minimize complications in anesthetic management. To the best of our knowlegde this is the first report in the literature which describes anesthesia for laparoscopic surgery in a SCI patient.