Induction of the 72-kD heat shock protein in xeroderma pigmentosum complementation group A fibroblasts.

In mammalian cells, 72-kD heat shock protein (HSP72) is the major stress-inducible protein that is thought to play a protective role against the various environmental stresses. In order to know the induction mechanism of HSP72, we examined the HSP72 in DNA repair-deficient xeroderma pigmentosum group A fibroblasts (XP2OSSV) and normal fibroblasts (WI38VA13) by the indirect immunofluorescence method using a monoclonal antibody specific for the inducible 72-kD protein. Heat-shock treatment of the same survival fraction (5% survival) induced HSP72 in xeroderma pigmentosum (XP) and normal cells. However, as compared with XP cells, normal cells showed the induction of HSP72 more rapidly and strongly. When XP and normal cells were irradiated with UVC at the same survival dose (10% survival), apparent induction of HSP72 was observed in both cell lines. In the case of UVC irradiation at the same dose (1.0 J/m2), though XP cells showed the induction of HSP72, HSP72 was not induced in normal cells. In both cell lines, heat-shock treatment caused more rapid induction of HSP72 than UV irradiation. These results suggest that the induction mechanism of HSP72 might be different between heat-shock treatment and UV irradiation. In addition, in the case of UV irradiation, the extent of DNA damage after DNA repair or the cell death might be involved in the induction of HSP72.

Increase in participation of vasoactive intestinal peptide in relaxation of the distal colon of Wistar rats with age.

Changes in participation of vasoactive intestinal peptide (VIP) in nonadrenergic noncholinergic (NANC) relaxation of longitudinal muscle of the distal colon with age were studied in 2- to 50-week-old Wistar rats in vitro. The extent of the VIP-mediated component of the relaxation induced by electrical field stimulation (EFS) was determined by the effect of VIP(10 - 28), a VIP receptor antagonist. In 2-week-old rats, the extent of the VIP-mediated component of the relaxation was scarce, about 10%, whereas the component gradually increase with age and reached the maximum extent 66% at 50-week-old. Since our previous results suggest that VIP induces NANC relaxation via activation of charybdotoxin (ChTx, a blocker of large conductance Ca(2+)-activated K(+) channel)-sensitive K(+) channels with concomitant slow hyperpolarization in the muscle cells, we next studied whether ChTx-sensitive component and slow hyperpolarization changes with age. Extent of ChTx-sensitive component of the relaxation increased with age, showing a very similar pattern to VIP-mediated one. EFS induced monophasic inhibitory junction potentials (i.j.ps) in longitudinal muscle cells of the distal colon of 2- and 4-week-old. EFS also induced biphasic i.j.ps in many longitudinal muscle cells of 8- and 50-week-old: rapid and subsequent slow hyperpolarization. A VIP receptor antagonist selectively inhibited the slow hyperpolarization. Exogenously added VIP induced no appreciable change in the membrane potential of longitudinal muscle cells of 2-week-old, whereas it induced slight slow hyperpolarization of the cell membrane in 4-week-old and magnitude of the hyperpolarization increased with age. On the other hand, relaxant response of the longitudinal muscle to exogenously added VIP was high in younger rats. The present results suggest that the role of VIP in mediating NANC relaxation of longitudinal muscle of the Wistar rat distal colon is very little at neonatal stage, but it increases with age.

Expression of prostaglandin endoperoxide synthase in rat brain.

Developmental and regional expression of prostaglandin endoperoxide synthase (PES) transcript was examined in the rat brain and in primary mixed cultures of neurons and glial cells from neonatal brain. Although the PES mRNA level in the brain was much lower than that in peripheral rat tissues such as lung, liver, spleen and kidney, a significant 3.0 kb band was detected in brain samples by Northern blot analysis. During development, PES mRNA was first detectable at postnatal day 7, and increased thereafter toward adulthood. The highest level of 3.0 kb PES mRNA was observed in the olfactory bulb, midbrain, and hypothalamus; and the lowest level in the hippocampus. In primary cultures of neonatal brain cells, the level of 3:0 kb transcript of PES transiently and dramatically increased about 30-fold on the third day after plating. Simultaneously, two cross-hybridizing signals were detected at 4.0 and 7.0 kb. This increase in PES mRNAs was completely inhibited by addition of cytosine-1-beta-D-arabinofuranoside. The induction of PES mRNA was in parallel with the increase in PES protein, as assessed by Western blot analysis. Immunostaining of cultured cells with anti-PES monoclonal antibody revealed that PES protein was induced mainly in neurons but not in glial cells. These results suggest that PES is expressed in the central nervous system at a low concentration under normal conditions, and that the neuronal cells possess an ability to express high levels of PES mRNA and protein.

Purification and some properties of rat intestinal ornithine decarboxylase.

Ornithine decarboxylase (ODC) was induced in rat small intestine by treatment with hypotonic solution in vitro and purified by two procedures, a conventional procedure and an immunoaffinity procedure. SDS-polyacrylamide gel electrophoresis showed that the molecular weight of the preparation purified by the immunoaffinity procedure (Mr = 53,000) was slightly larger than that of the preparation obtained by the conventional procedure (Mr = 52,000). Values for the Km for L-ornithine (0.1 mM), the isoelectric point (5.4), and the final specific activity (5.1-5.5 x 10(5) nmol CO2/mg protein/30 min) of the two preparations were similar to those reported for the rat liver ODC. Addition of a protease inhibitor (limabean trypsin inhibitor) to the crude extract prevented the appearance of the smaller enzyme (Mr = 52,000) obtained by the conventional purification procedure. Our result indicates that the large enzyme is native ODC and the smaller one is a partial proteolysis product of native ODC.

Occurrence of okadaic acid-producing Prorocentrum lima on the Sanriku coast, northern Japan.

Prorocentrum lima was found to be distributed on the surface of the algae, Sargassum confusum and Carpopeltis flabellata collected at the Sanriku coast, northern Japan. Chemical analysis of cultured cells revealed that Sanriku strains of P. lima produce okadaic acid, a toxin responsible for diarrhetic shellfish poisoning. The Sanriku strain grew well in T1 medium at 15 degrees C at which tropical strains do not grow, indicating that it is a local strain which adapts to cooler environments.

8-methoxypsoralen plus UVA induces the 72 kDa heat shock protein in organ-cultured normal human skin.

The proteins induced by heat and other stressors, called heat shock proteins (HSP) or stress proteins, are considered to play a general role in protection from cellular injury. Exposure to UVA (320-400 nm) following application of 8-methoxypsoralen (8-MOP), termed PUVA is commonly used in the field of dermatology. In order to understand the induction of HSP in PUVA-treated human skin, indirect immunofluorescence using a monoclonal antibody specific for the 72 kDa HSP (HSP 72) was carried out in organ-cultured normal human skin that was treated with PUVA. When the organ-cultured skin was treated at 37 degrees for 1 h with 8-MOP at a final concentration of 10 or 100 micrograms/mL and exposed to UVA (51.3 kJ/m2), nuclear immunofluorescence of HSP 72 was detected in the epidermal cells 12 h after UVA irradiation. In contrast, the induction of HSP 72 was not detected either by UVA irradiation or 8-MOP treatment. These results suggest that PUVA treatment is one of the stressors for human skin, and DNA damage caused by PUVA induces HSP 72.

Induction and repair of UVB-induced cyclobutane pyrimidine dimers and (6-4) photoproducts in organ-cultured normal human skin.

To examine the induction and repair of UV-induced DNA damage, indirect immunofluorescence was performed on UVB-irradiated organ-cultured normal human skin using monoclonal antibodies specific for either cyclobutane pyrimidine dimers or (6-4) photoproducts. Nuclear immunofluorescence of cyclobutane pyrimidine dimers and (6-4) photoproducts were observed in a dose-dependent manner after UVB irradiation. The intensity of nuclear immunofluorescence of the upper epidermal layers was stronger and clearer than that of the lower epidermal layers. DNA repair time-course studies showed that both types of DNA damage could be repaired within 24 h after UVB irradiation.

Protein synthesis by intracellular symbionts in two closely interrelated aphid species.

An aphid endosymbiont in vivo synthesizes symbionin almost exclusively which is not produced in vitro by the same symbiont. While symbionin produced by the endosymbiont of the pea aphid is an acidic protein with a molecular weight of 63,000, that by the symbiont of the kondo aphid, the closest relative to the former, is a distinct, less acidic, molecule. While the two endosymbionts in vivo in old insects synthesize about 11 protein species in common, they produce many different proteins when incubated extracellularly.

Role of substance P in experimental allergic conjunctivitis in guinea pigs.

The role of substance P in experimental allergic conjunctivitis induced by egg albumin was investigated with guinea pigs. Increase in vascular permeability of the conjunctiva induced by antigen was significantly inhibited after repeated application of capsaicin. Substance P contents in the conjunctiva of guinea pig were decreased by topical instillation of antigen to the eyes, suggesting that substance P was released from the conjunctiva due to antigen-antibody reaction. Moreover, subconjunctival injection of substance P resulted in a dose-related conjunctivitis, and vascular permeability in the conjunctiva was also increased by substance P. In substance P-induced conjunctivitis, a significant edema was observed in the bulbar and palpebral conjunctiva, but no hyperemia was noted in all instances. Histamine contents of the conjunctiva and tears were not influenced by subconjunctival injection of substance P. However, topical application of antigen and subconjunctival injection of compound 48/80 caused a significant decrease in histamine content, and content of tear was increased by both treatments. An increase in vascular permeability induced by antigen application was significantly inhibited by intravenous injection of FK-888, which is a specific and potent NK1 receptor antagonist. From these results, it is suggested that substance P is responsible for allergic conjunctivitis to some extent, and the conjunctival hyperpermeability induced by substance P occurs through NK1 receptor on the blood vessels, rather than by the direct action on the conjunctival mast cells during allergic conjunctival reactions.

Mediators of nonadrenergic, noncholinergic relaxation in Sprague Dawley rat intestine: comparison with the mediators of other strains.

Participation of nitric oxide, vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP) in nonadrenergic, noncholinergic (NANC) relaxation of longitudinal muscle of various intestinal regions in Sprague Dawley rats (8-week-old) was studied in vitro. Nitric oxide was suggested to participate in NANC relaxation of every intestinal region studied. But the participation was partial and its extent varied among the regions: significant in the proximal colon and rectum, and moderate in the jejunum, ileum and distal colon. Participation of PACAP in NANC relaxation was suggested only in the distal colon, while that of VIP was not detected in any of regions. Results obtained in the present study indicate that extent of participation of nitric oxide in NANC relaxation in Sprague Dawley rat intestine is more significant than those of other strains, Wistar and Wistar-ST.

Lower third molar infection with purulent discharge through the external auditory meatus. Case report and review of literature.

This manuscript reports an uncommon case of inferior third molar facial abscess with purulent secretion drainage through the left external acoustic meatus. The patient's left external acoustic meatus was filled with a purulent secretion observed on a CT scan. He underwent surgery to drain the facial abscess. Despite facial abscesses being routine occurrences, the literature does not contain many case reports of odontogenic facial abscesses with drainage via the external acoustic meatus. These situations occur in two possible ways: multiple fissures in the anterior wall of the cartilaginous portion of the external acoustic meatus; and congenital defects that are occasionally present in the anterior-superior aspect of the external acoustic meatus, known as the foramen of Huschke, which allow communication between the external acoustic meatus and mandibular fossa. These defects may also predispose the patient to the spread of the infection or tumour from the external auditory canal to the infratemporal fossa and vice versa. No otological sequelae were observed in this case. The authors conclude that the hypothesis of bone malformation cannot be excluded, and affirm that any facial abscess requires appropriate and immediate treatment for adequate resolution, by removing the causal factor and providing systemic support.