Hedgehog inhibitors from Artocarpus communis and Hyptis suaveolens.

The hedgehog (Hh) signaling pathway plays crucial roles in cell maintenance and proliferation during embryonic development. Naturally occurring Hh inhibitors were isolated from Artocarpus communis and Hyptis suaveolens using our previously constructed cell-based assay system. Bioactivity guided fractionation led to the isolation of 15 compounds, including seven new compounds (4, 5, 6, 7, and 9-11). The isolated compounds showed cytotoxicity against a cancer cell line (PANC1) in which Hh signaling was abnormally activated. Several compounds (12-14; GLI1 transcriptional inhibition IC50=7.6, 4.7, and 4.0 µM, respectively) inhibited Hh related protein (BCL2) expression. Moreover, compounds 1, 12, and 13 disrupted GLI1 and DNA complex formation.

Activation of Master Autophagy Regulator TFEB During Systemic LPS Administration in the Cornea.

The involvement of autophagy in the cornea during the systemic inflammatory response elicited by intravenous administration of lipopolysaccharide (LPS) was investigated. Eight-week-old male Sprague-Dawley rats were injected i.v. with 15 mg/kg body weight LPS. RC4 rabbit corneal keratocytes were also used and treated with 100 ng/mL of tumor necrosis factor α (TNFα) and/or cycloheximide (CHX). The nuclear translocation of transcription factor EB (TFEB), the master transcriptional regulator for autophagy, was observed after LPS administration in the corneal epithelium. Induction of autophagy-related proteins was observed in the cornea after LPS administration, as well as in RC4 cells after treatment with TNFα. Administration of trehalose, an inducer of TFEB, mitigated RC4 cell death caused by TNFα/CHX. These results demonstrate the importance of TFEB activation in cellular defense against the systemic inflammatory response in the cornea.

Physalin H from Solanum nigrum as an Hh signaling inhibitor blocks GLI1-DNA-complex formation.

Hedgehog (Hh) signaling plays an important role in embryonic development, cell maintenance and cell proliferation. Moreover, Hh signaling contributes to the growth of cancer cells. Physalins are highly oxidized natural products with a complex structure. Physalins (1-7) were isolated from Solanum nigrum (Solanaceae) collected in Bangladesh by using our cell-based assay. The isolated physalins included the previously reported Hh inhibitors 5 and 6. Compounds 1 and 4 showed strong inhibition of GLI1 transcriptional activity, and exhibited cytotoxicity against cancer cell lines with an aberrant activation of Hh signaling. Compound 1 inhibited the production of the Hh-related proteins patched (PTCH) and BCL2. Analysis of the structures of different physalins showed that the left part of the physalins was important for Hh inhibitory activity. Interestingly, physalin H (1) disrupted GLI1 binding to its DNA binding domain, while the weak inhibitor physalin G (2) did not show inhibition of GLI1-DNA complex formation.

Hh signaling inhibitors from Vitex negundo; naturally occurring inhibitors of the GLI1-DNA complex.

The hedgehog (Hh) signaling pathway has crucial roles in embryonic development, cell maintenance and proliferation, and is also known to contribute to cancer cell growth. New naturally occurring Hh inhibitors (1, 7 and 9) were isolated from Vitex negundo using our previously constructed cell-based assay. Bioactivity guided isolation provided 9 natural compounds including a new diterpene, nishindanol (9). Compounds 7 and 9 showed cytotoxicity against cancer cell lines in which Hh signaling was aberrantly activated. Vitetrifolin D (7; GLI1 transcriptional inhibition IC50 = 20.2 µM) showed inhibition of Hh related protein (PTCH and BCL2) production. Interestingly, the constructed electrophoresis mobility shift assay revealed that vitetrifolin D (7) disrupted GLI1 binding on its DNA binding domain. epi-Sclareol (8; inactive), possessing a similar structure to 7, did not show inhibition of GLI1­DNA complex formation. This is the first example of naturally occurring inhibitors of GLI1­DNA complex formation.

Autopsy findings of a patient with rapidly progressive massive ascites caused by alcoholic cirrhosis.

A 54-year-old man, who lived alone, was hospitalized due to rapid deterioration of the general condition over a three-week period caused by alcoholic cirrhosis. One month after he left hospital, he was found dead in his house by his friend. Three days before he was found dead, he had met his friend and seemed to be in poor condition. Autopsy was conducted by a medical examiner to clarify the cause of death. Externally, signs of severe jaundice were apparent over the whole body, along with extensive abdominal swelling and edema of the extremities. Autopsy findings demonstrated that the abdominal cavity contained an amount of massive turbid and slight pale reddish brown ascites (23 l). There were no findings of severe peritoneal inflammation. The liver (650 g) was elastic hard and had a micro-nodular surface, which showed severe atrophy. Microscopic examination of the liver showed clear pseudolobule with severe fibrosis in the stroma. There were no significant changes in the heart or brain. The stomach was empty and only a slight amount of intestinal contents. There was no ethanol detected in the blood or urine. The direct cause of his death was circulatory dysfunction due to massive accumulation of the ascites. The reasons for the massive ascites accumulation over 20 l in this case were (1) that he had no serious complications other than ascites; and (2) he did not have any medical treatment just before his death.

Death due to blood transfusion-induced anaphylactic shock: A case report.

In medical practice, many clinical accidents due to blood transfusion reactions have been reported, among which, nonhemolytic transfusion reactions (NHTRs) have been mainly reported in recent years. NHTRs induce reactions such as anaphylactic shock and transfusion-related acute lung injury (TRALI); however, few forensic autopsy case reports with blood transfusion reactions including anaphylactic shock have been published. A marker for anaphylactic shock is serum tryptase, which indicates systematic mast cell activation in living patients. In forensic medicine, serum tryptase has been used in the postmortem diagnosis of anaphylactic shock. In this autopsy case report, the blood tryptase level was elevated to 49.0 ng/mL (reference standard level <13.5 ng/mL). When considered comprehensively with autopsy findings and blood typing, we concluded that this patient was suspected to have suffered from anaphylactic shock as a result of blood transfusion.

A fatal case of air-bag-mediated liver injury in an unrestrained driver.

A middle-aged man, who drove a small vehicle while drunken and not wearing a seat belt, crashed head on into a pier wall at a high speed. The rapid air-bag deployment caused abdominal compression of the driver, who was found dead 2 m away from the car. There were facial front-glass injuries and multiple rib fractures, but no head injury. Autopsy disclosed liver laceration with intra-abdominal hemorrhage (2,000 mL), resulting in hemorrhagic shock. This is the first autopsy report of liver laceration caused by the rapid expansion of an air-bag during an automobile crash.