Impact of adrenomedullin on dextran sulfate sodium-induced inflammatory colitis in mice: insights from in vitro and in vivo experimental studies.

BACKGROUND: Although adrenomedullin (AM) is known to ameliorate inflammatory processes, few data exist regarding the effect of AM on inflammatory colitis. Therefore, we examined the effect of AM on inflammatory response in vitro and in vivo colitis model. METHODS: In mice experimental colitis induced by 3% dextran sulfate sodium (DSS) in drinking water for 7 days, AM with 225-900 µg/kg in 0.5 ml of saline or saline alone were given intraperitoneally once a day. In the in vitro experiment, we determined the cytokine response in THP-1 cell activated by lipopolysaccharide with or without AM of 10 nM. Additionally, we performed wound healing assay in Caco-2 cell interfered by DSS with or without AM of 100 nM. RESULTS: In the colitis model, AM significantly reduced the disease activity index, histological score, and local production of inflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 in accordance with reduction of serum amyloid A levels. Secretion of TNF-α in lipopolysaccharide-stimulated THP-1 cells was significantly reduced in the presence of AM. The distance of wound healing interfered by 0.25% DSS was significantly improved in the presence of AM of 100 nM. CONCLUSIONS: These results demonstrate that AM could ameliorate DSS-induced experimental colitis possibly through suppression of systemic and local production of cytokines such as TNF-α, associated with acceleration of ulcer reepithelialization and colon tissue regeneration.

[Present situation of crime in Japan].

Recently, it has been said that public safety has changed for the worse in Japan. After a peak in 1994, the total number of crimes gradually decreased, but it still remains at a higher level when compared to that in 1989. In addition to the persistent increase in the number of child abuse and drug abuse/dependence cases as well as stalking, indiscriminate crime by an unknown assailant is now increasing in Japan, particularly this year. The present criminal tendency seems to be based on the rapid change from an equal to unequal societal status, the ease of access to personal computers/the Internet, and the increase of crime committed by foreigners.

Involvement of human blood arylesterases and liver microsomal carboxylesterases in nafamostat hydrolysis.

Metabolism of nafamostat, a clinically used serine protease inhibitor, was investigated with human blood and liver enzyme sources. All the enzyme sources examined (whole blood, erythrocytes, plasma and liver microsomes) showed nafamostat hydrolytic activity. V(max) and K(m) values for the nafamostat hydrolysis in erythrocytes were 278 nmol/min/mL blood fraction and 628 microM; those in plasma were 160 nmol/min/mL blood fraction and 8890 microM, respectively. Human liver microsomes exhibited a V(max) value of 26.9 nmol/min/mg protein and a K(m) value of 1790 microM. Hydrolytic activity of the erythrocytes and plasma was inhibited by 5, 5'-dithiobis(2-nitrobenzoic acid), an arylesterase inhibitor, in a concentration-dependent manner. In contrast, little or no suppression of these activities was seen with phenylmethylsulfonyl fluoride (PMSF), diisopropyl fluorophosphate (DFP), bis(p-nitrophenyl)phosphate (BNPP), BW284C51 and ethopropazine. The liver microsomal activity was markedly inhibited by PMSF, DFP and BNPP, indicating that carboxylesterase was involved in the nafamostat hydrolysis. Human carboxylesterase 2 expressed in COS-1 cells was capable of hydrolyzing nafamostat at 10 and 100 microM, whereas recombinant carboxylesterase 1 showed significant activity only at a higher substrate concentration (100 microM). The nafamostat hydrolysis in 18 human liver microsomes correlated with aspirin hydrolytic activity specific for carboxylesterase 2 (r=0.815, p<0.01) but not with imidapril hydrolysis catalyzed by carboxylesterase 1 (r=0.156, p=0.54). These results suggest that human arylesterases and carboxylesterase 2 may be predominantly responsible for the metabolism of nafamostat in the blood and liver, respectively.

Fatal heat injury incident in a private home for elderly people: a case report.

This report presents a case in which an 84-year-old elderly female resident of a private home for the aged was fatally injured by a young male member of staff. The deceased suffered from senile dementia, and died from heat shock due to prolonged exposure to an electric fan heater. The heater had been placed immediately in front of the woman's body for approximately 2.5 hours, and the deceased could not escape from the heat by herself. Major autopsy findings were marked second degree burns (predominantly) and smaller areas of first degree burns observed on the anterior body surface (chiefly the face, lower abdomen, forearm and thigh). The burns area exceeded one-third of the entire body surface. Only a small amount of dark red blood (36g) remained in the heart cavity: this was fluid but remarkably condensed. Petechial bleeding was observed in areas such as the palpebral conjunctivae and renal pelvic mucosae. This case generated great public attention as the assailant, who had been employed exclusively for three nights' duty per week, had been regarded as very diligent and kind; inflicting such a lethal injury appeared out of character. Several important issues related to managing residential care facilities for the elderly (in particular, the stress associated with night duty caring for the aged with senile dementia, and the mental health care of the staff themselves) were disclosed and are discussed herein from a medico-legal viewpoint.

Accelerated wound healing in tumor necrosis factor receptor p55-deficient mice with reduced leukocyte infiltration.

To clarify biological roles of tumor necrosis factor receptor p55 (TNF-Rp55) -mediated signals in wound healing, skin excisions were prepared in BALB/c (WT) and TNF-Rp55-deficient (KO) mice. In WT mice, the wound area was reduced to 50% of the original area 6 days after injury, with angiogenesis and collagen accumulation. Histopathologically, reepithelialization rate was approximately 80% 6 days. Myeloperoxidase activity and macrophage recruitment were the most evident 1 and 6 days after injury, respectively. Gene expression of adhesion molecules, interleukin 1alpha (IL-1alpha), IL-1beta, monocyte chemoattractant protein 1, macrophage inflammatory protein 1alpha (MIP-1alpha), MIP-2, transforming growth factor beta1 (TGF-beta1) connective tissue growth factor (CTGF), vascular endothelial growth factor (VEGF), Flt-1, and Flk-1 was enhanced at the wound site. In KO mice, an enhancement in angiogenesis, collagen content, and reepithelialization was accelerated with the increased gene expression of TGF-beta1, CTGF, VEGF, Flt-1, and Flk-1 at the wound sites, resulting in accelerated wound healing compared with WT mice. In contrast, leukocyte infiltration, mRNA expression of adhesion molecules, and cytokines were significantly reduced in KO mice. These observations suggest that TNF-Rp55-mediated signals have some role in promoting leukocyte infiltration at the wound site and negatively affect wound healing, probably by reducing angiogenesis and collagen accumulation.

A pivotal involvement of IFN-gamma in the pathogenesis of acetaminophen-induced acute liver injury.

In wild-type BALB/c mice, i.p. administration of acetaminophen (APAP; 750 mg/kg) induced intrahepatic IFN-gamma mRNA expression and a marked increase in serum transaminase levels, leading to acute lethality of approximately 45%. Histopathological examination showed centrilobular hepatic necrosis with leukocyte infiltration and a large number of apoptotic hepatocytes 10 and 24 h after APAP challenge. mRNA expression of intercellular adhesion molecule 1, vascular cell adhesion molecule 1, interleukin (IL) 1alpha, IL-1beta, IL-6, tumor necrosis factor alpha, monocyte chemoattractant protein 1, macrophage inflammatory protein (MIP) 1alpha, MIP-2, KC, IP-10, Mig, Fas, and inducible nitric oxide synthase was enhanced in the liver of wild-type mice injected with APAP. To clarify the role of IFN-gamma in this process, IFN-gamma-deficient mice were treated in the same manner. All IFN-gamma-deficient mice survived with reduced serum transaminase elevation and attenuated hepatic necrosis, leukocyte infiltration, and hepatocyte apoptosis. The gene expression of all molecules was significantly attenuated in IFN-gamma-deficient mice. Administration of an anti-IFN-gamma neutralizing antibody even 2 or 8 h after APAP challenge to wild-type mice alleviated APAP-induced liver injury, and all mice survived. Thus, IFN-gamma is responsible for APAP-induced liver injury by mediating leukocyte infiltration, hepatocyte apoptosis, and NO production as well as cytokine and chemokine production. Moreover, immunoneutralization of IFN-gamma may be therapeutically effective for developing APAP-induced liver injury.

Major cytochrome P450 enzymes responsible for microsomal aldehyde oxygenation of 11-oxo-Delta8-tetrahydrocannabinol and 9-anthraldehyde in human liver.

Hepatic microsomes from human liver catalyzed oxidation of the allyl aldehydes such as 11-oxo-Delta(8)-tetrahydrocannabinol and 9-anthraldehyde to the corresponding carboxylic acid metabolites. The oxygenation mechanism was confirmed by GC-MS that molecular oxygen was exclusively incorporated into Delta(8)-tetrahydrocannabinol-11-oic acid and 9-anthracene carboxylic acid formed under oxygen-18 gas. The microsomal aldehyde oxygenase (named MALDO) activities of 11-oxo-Delta(8)-tetrahydrocannabinol and 9-anthraldehyde were significantly inhibited by the antibody against CYP2C and CYP3A, respectively. MALDO activity for 11-oxo-Delta(8)-tetrahydrocannabinol was significantly inhibited by sulfaphenazole whereas that for 9-anthraldehyde was markedly inhibited by troleandomycin, but not by sulfaphenazole. CYP2C9 expressed in human B-lymphoblastoid cells catalyzed efficiently the MALDO activity for 11-oxo-Delta(8)-tetrahydrocannabinol (10.1 nmol/min/nmol P450), while the catalytic activities of other human CYPs expressed in the cells were lesser extents. In MALDO activity for 9-anthraldehyde, CYP3A4 expressed in the cells had the highest catalytic activity (7.72 nmol/min/nmol P450). These results indicate that CYP2C9 and CYP3A4 are major enzymes responsible for the MALDO activity in human liver for 11-oxo-Delta(8)-tetrahydrocannabinol and 9-anthraldehyde, respectively.

Air bag injuries--a literature review in consideration of demands in forensic autopsies.

Air bags have been implicated in saving lives and reducing morbidity associated with motor vehicle crashes since their introduction in the mid-1970s. However, there is increasing evidence showing that air bags can be a source of injury and even death in certain circumstances. As the number of air bag-equipped vehicles increases, air bag-related injuries have occurred more frequently. Thus, a greater awareness of air bag-related injuries is required in forensic autopsies. Here, we review thoroughly the literature concerning air bag-related injuries with special regard to their nature and causative mechanisms, and summarize air bag-related injuries observed in adults, children and infants.

The gene expression of cytokines and chemokines induced by tourniquet shock in mice.

Traumatic shock is one of the major fields in forensic pathology, but its mechanism remains elusive from the pathophysiological aspects. Tourniquet shock has been established as one of the animal models of traumatic shock, and we examined the gene expression of cytokines and chemokines in the lung and liver in tourniquet shock using mice. Tourniquet was conducted by the application of elastic bands with five turns at both the thighs as high as possible for 2 h, followed by reperfusion. In this procedure, more than 90% mice died within 48 h after reperfusion. Serum hepatic transaminase and hematocrit values significantly increased at 2 h after reperfusion, and their elevation was still evident after 10 h. Histopathologically, hemorrhages, congestion and leukocyte recruitment were observed in the lung and liver specimens after 6 h of reperfusion. Immunohistochemical analysis with anti-myeloperoxidase antibody demonstrated a massive neutrophil infiltration in the lung and liver at 2 h or more after reperfusion. RT-PCR analyses demonstrated that the gene expression of interleukin-1beta, tumor necrosis factor-alpha, monocytes chemoattractant protein-1, macrophage inflammatory protein (MIP)-1alpha, MIP-2, KC and vascular endothelial adhesion molecule-1 was most enhanced in the lung and liver at 2 h after reperfusion. Thus, the gene expression of cytokines and chemokines is presumed to be closely related with the onset of tourniquet shock. From the forensic aspects, these cytokines and chemokines are considered to be useful markers for the early diagnosis of tourniquet shock.

Tracheal injury added to cervical bone destruction due to the impact of hitting the water surface: four immersed adult bodies.

In the forensic examinations of cases of falling, two properties of the water surface, namely its nature as a hard, flat object and as a soft and ungraspable substance must be appreciated. Namely, at the moment of impact, the water surface exerts a greater resistance against relatively broad areas like the head, face and trunk than against the extremities that have a small area. Therefore, total resistance against the whole body would promote flexure. We experienced 72 autopsy cases of immersed bodies during a 4-year period. The cause of death for 64 of these with or without cervical vertebra fracture was drowning. In these cases, the various heights of the falls could often be estimated at the scene. A characteristic pattern of cervical injury with involvement of hyoid bone and thyroid cartilage in addition to cervical vertebra fracture plus rare involvement of the trachea was identified. When a fall from a relatively low height is broken by the water surface, to a certain degree physical findings that differ from those seen in falls to the ground from extreme heights are left mediated by different underlying mechanisms.

Fatal and non-fatal cases of lime sulfide exposure and pathogenetic mechanisms underlying pancreatic injury: Case reports with an animal experiment.

Lime sulfide poisoning by the oral route is rarely encountered in the practice of forensic science, whereas hydrogen sulfide poisoning is seen frequently. We report here two cases of fatal lime sulfide poisoning with several related cases and in addition induced histological damage with acute inflammation in animal models under at similar concentrations. We also evaluated sulfide and thiosulfate concentrations and speculated as to the cause of pancreatic damage in these cases.

Sudden death of a young male with previously undiagnosed autosomal dominant polycystic kidney disease (ADPKD).

Autosomal dominant polycystic kidney disease (ADPKD) is one of the best known genetic diseases. However, in only very rare cases does it present as an abnormal death followed by clarification of its genetic background. We experienced a case in which ADPKD first became evident from the results of forensic autopsy, on the basis of which all potentially affected family members were offered genetic and other medical examinations. In this way, forensic medicine was able not only to determine the cause of death but to contribute to preventive medicine as well.

Impairment of skin wound healing in beta-1,4-galactosyltransferase-deficient mice with reduced leukocyte recruitment.

Cell-surface carbohydrate chains are known to contribute to cell migration, interactions, and proliferation, but their roles in skin wound healing have not been evaluated. We examined the biological roles of beta4-galactosylated carbohydrate chains in skin wound healing using mutant mice that lack beta-1,4-galactosyltransferase-I (beta4GalT-I), which is responsible for the biosynthesis of the type 2 chain in N-glycans and the core 2 branch in O-glycans. beta4GalT-I-deficient mice showed significantly delayed wound healing with reduced re-epithelialization, collagen synthesis, and angiogenesis, compared with control mice. Neutrophil and macrophage recruitment at wound sites was also impaired in these mice probably because of selectin-ligand deficiency. In accordance with the reduced leukocyte infiltration, the expression levels of macrophage-derived chemokines, transforming growth factor-beta1, and vascular endothelial growth factor were all reduced in beta4GalT-I(-/-) mice. These results demonstrate that beta4-galactosylated carbohydrate chains play a critical role in skin wound healing by mediating leukocyte infiltration and epidermal cell growth, which affects the production of chemokines and growth factors. This study introduces a suitable mouse model for investigating the molecular mechanisms of skin wound healing and is the first report showing that carbohydrate chains have a strong influence on skin wound healing.