Takaaki Matsumoto

TGF-β1 mediates 70-kDa heat shock protein induction due to ultraviolet irradiation in human skin fibroblasts

Abstract Ultraviolet B (UVB) alters the expression of heat shock protein 70 (HSP70) in cultured fibroblast cells derived from human skin. However, the nature of the signal transduction pathway remains to be determined. Transforming growth factor-β (TGF-β) has a large variety of biological functions, including cell growth control, modulation of inflammation and immunoregulation. In this study, we examined whether TGF-β is associated with the process of HSP70 expression induced by UVB irradiation. The constitutive expression of TGF-β1 mRNA and HSP70 expression in human skin fibroblast cells were detected using reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis. The results indicate that: (1) UVB irradiation stimulates HSP70 expression in a dose- and time-dependent manner, (2) constitutive expression of TGF-β1 mRNA is detected after UVB irradiation, the level of which peaks at 4 h after 10 mJ cm–2 of UVB irradiation, (3) HSP70 expression is induced by TGF-β1 without UVB irradiation, and (4) HSP70 expression induction with UVB irradiation is inhibited by preincubation of the cells with the anti-TGF-β type II receptor antibody. Our results suggest that HSP70 expression induced by UVB involves the autocrine signalling of TGF-β production.

Irregular activation of individual sweat glands in human sole observed by a videomicroscopy

Sweat secretion from individual sweat glands on the human sole was observed in four male subjects by using a videomicroscope and correlated with sudomotor neural activity recorded from the tibial nerve by means of microneurography. Individual sweat glands could be distinguished as active, less active and inactive according to the incidence of sweat secretion during spontaneous sweating. The threshold amplitude of the sudomotor burst necessary for sweat secretion varied from gland to gland. The number of sweat secretion was significantly related to the threshold amplitude. Sweat glands often failed to produce sweat secretion even when a suprathreshold burst occurred: only 46.1+/-3.8% (mean +/- S.E.M.) of the suprathreshold bursts elicited sweat secretion. Failure of the sweat secretion tended to appear after several bursts occurred consecutively with short intervals. In spite of the variability in sweat gland activity, the number of sweat glands recruited was linearly related to the amplitude of the sudomotor burst (P < 0.001). Thus, although sweat secretion from each sweat gland depends primarily on the intensity of sudomotor neural activity. the activity of each sweat gland may fluctuate temporally as the result of irregular activation of sudomotor fibers and possibly some intrinsic factors of the gland.

Hyperthermic Enhancement of the Apoptotic and Antiproliferative Activities of Paclitaxel

The antineoplastic agent paclitaxel (PTX), a microtubule-stabilizing agent, is known to arrest cell cycle progression and induce apoptosis. Mild hyperthermia (HT) also disrupts the microtubule system and triggers apoptosis. We therefore investigated whether concurrent exposure of murine breast cancer cells to 10 µmol/l PTX and 43°C HT will promote improved anticancer effects. To do this, we exposed FM3A murine cancer cells to: (1) 10 µmol/l PTX for 1 h at 37°C followed by exposure at 43°C HT for 1 h; (2) 10 µmol/l PTX at 37°C for 2 h; (3) 37°C for 1 h followed by 43°C HT for 1 h, and (4) untreated cells at 37°C for 2 h which served as the control. Treatment No. 1 resulted in an enhanced cell cycle arrest, apoptosis and cytotoxicity. Exposure to 43°C HT alone or 10 µmol/l PTX alone induced lesser apoptosis and cytotoxicity than the two treatments concurrently applied. The apoptotic cell death occurred in a time-dependent manner as follows: (1) concurrently applied 43°C HT and 10 µmol/l PTX (5.6 ± 0.5, 16.5 ± 2 and 27.6 ± 1%); (2) 43°C HT alone (4.3 ± 1, 6.6 ± 0.3 and 12.7 ± 1%) and (3) 10 µmol/l PTX alone (4.4 ± 0.3, 8.6 ± 1 and 12.8 ± 1%) at 1, 6 and 24 h postexposure respectively compared to control of 2.0%. These data indicate that while both HT and PTX can individually induce apoptosis and antiproliferation in FM3A cancer cells, they may offer synergistic benefits when used concurrently.

Contribution of nitric oxide to adaptation of tibetan sheep to high altitude

We examined the effects of endogenous nitric oxide synthase (NOS) inhibition on pulmonary hemodynamics in awake sheep living at low and high altitudes to evaluate the role of NO in adaptation to an hypoxic environment. Unanaesthetized male sheep in three places--Matsumoto, Japan (680 m above sea level), Xing, China (2300 m) and Maxin, China (3750 m)--were prepared for measurements of pulmonary artery (Ppa) and pulmonary vascular resistance (PVR) before and after the NOS inhibition. The non-selective NOS inhibitor, Nw-nitro-l-argine (NLA, 20 mg/kg) was used. Baseline Ppa became elevated with an increase in altitude. After NLA administration, PVR significantly increased in animals of all groups. However, the increase in PVR after NLA in tibetan sheep at 3750 m was significantly higher than those in other groups. We conclude that augmented endogenous NO production may contribute to regulating the pulmonary vascular tone in tibetan sheep (3750 m) adapted to high altitude.

Low and high frequency acupuncture stimulation inhibits mental stress-induced sweating in humans via different mechanisms.

The effects of acupuncture stimulation at 5 Hz and 100 Hz on mental stress-induced sweating were analyzed, and the mechanisms involved were examined using the rate of sweat expulsion as an estimate of central sudomotor outflow. Mental arithmetic was imposed on 25 young healthy volunteers for 2 min before, during and after the stimulation. Acupuncture stimulation was delivered to either the Zusanli (leg) or Hegu (hand) acupoint, and the sweat rate was measured quantitatively during mental arithmetic on the palm or the sole, respectively. When stimulation at 5 Hz was applied to the Zusanli acupoint, the palmar sweat rate (paSR), rate of sweat expulsion (Fsw) and paSR/Fsw were reduced during the stimulation, whereas when it was applied to the Hegu acupoint, plantar SR (plSR) and Fsw were reduced, but plSR/Fsw was not altered. When stimulation at 100 Hz was applied to Zusanli, paSR and paSR/Fsw were reduced, but Fsw was unchanged whereas when it was applied to Hegu, neither plSR, Fsw nor plSR/Fsw was altered. The results suggest that acupuncture stimulation at 5 Hz affects both the supraspinal rhythm-generating mechanism and the mechanisms situated below (probably the spinal cord), whereas stimulation at 100 Hz only affects the mechanisms below the rhythm-generating mechanism. Thus, acupuncture stimulation at 5 Hz and at 100 Hz may reduce mental stress-induced sweating through different mechanisms.

Immunization with Trichinella spiralis Korean Isolate Larval Excretory–Secretory Antigen Induces Protection and Lymphocyte Subset Changes in Rats

Protection and immune responses were studied in rats immunized with Trichinella spiralis muscle stage larval excretory–secretory antigen (ES Ag) without adjuvant. Protection was assessed by the degree of adult worm burden and the yield of muscle (diaphragmatic) larvae after challenge infection with live larvae. Lymphocyte subsets were identified by flow cytometry in the spleen and peripheral blood. Cytokine production and specific IgG, IgG1 and IgG2a antibody responses were measured. Immunization with ES Ag produced highly significant protection against adult stages (98.4%) and muscle larvae (82.9%). Th2 type cytokines (IL‐10, IL‐4) were predominant. Anti‐muscle stage larval ES Ag antibody was significantly elevated in the order IgG2a > IgG1 > IgG on the 2nd day after final immunization and on the 7th day after challenge infection. Expression of CD4+ and the CD4+/CD8+ ratio from spleen and blood were significantly increased compared to the control. These results demonstrate that immunization with T. spiralis antigen can elicit robust immune response, resulting in complete protection against infective larvae, and this protection can be achieved without the use of any adjuvant.

The Role of RANTES in a Murine Model of Food Allergy

Food allergy is an important and common health issue, and there is a need to identify and characterize the sensitizing mechanisms. One of the common causes of food allergy is ovalbumin (OVA), a dietary antigen from eggs. We hypothesized that OVA‐induced food allergy in the gut involves the activation of the chemokine regulated on activation, normal T cell expressed and secreted (RANTES), which then recruits eosinophils to lesioned tissue. The purpose of this study was to clarify whether RANTES expression correlates with eosinophil infiltration in the gut of OVA‐sensitized BALB/c mice in response to oral OVA challenge. BALB/c mice were immunized with OVA 1 µg and sensitized after 2 weeks by intragastric administration of OVA. Sensitization to the oral OVA challenge was analyzed by examining eosinophil infiltration into the gut tissue (immunohistochemistry), mucosal eosinophil cationic protein (ECP) concentration, and RANTES mRNA expression (reverse‐transcriptase polymerase chain reaction and Southern blotting) at 3, 6, 12, and 24 h after the challenge. There was marked edema of the intestinal villi, and eosinophil infiltration to the lamina propria peaked at 6 h in OVA‐sensitized mice. RANTES mRNA expression peaked at 3 h and 6 h and declined thereafter. The expression of RANTES mRNA in the allergic mice was much higher than in the nonallergic, normal, or unsensitized control mice. Tissue eosinophilia and intestinal ECP levels were significantly correlated with the RANTES mRNA level. We conclude that RANTES may play a central role in the pathogenesis of food‐mediated gastrointestinal allergy.

Relationships Between IgG, IgM, IgE and Resistance to Reinfection During the Early Phase of Infection with Clonorchis sinensis in Rats

An enzyme linked immunosorbent assay (ELISA) was used to study the correlation between the levels of IgG, IgM and IgE immunoglobulin isotypes and resistance to re‐infection in rats during the first month of infection with Clonorchis sinensis. Rats were infected with Clonorchis sinensis (primary infection), and then treated with praziquantel on the 1st, 3rd, 7th, 14th and 28th day post infection (p.i.). To measure resistance, rats were re‐infected with C. sinensis (secondary infection), 2 weeks after the treatment and worms were recovered 4 weeks later. During the primary infection, significantly increased levels of IgG isotype were observed on days 14 and 28 p.i. (P < 0.001) and IgM levels were significantly increased on 3rd and 28th day (P < 0.001). During the secondary infection, significantly increased levels of IgG isotype were found from 3rd to 28th day and IgE isotype on 7th and 14th day (P < 0.01) while significant levels of IgM were found on the 3rd and 28th day (P < 0.05). Furthermore, significant differences of worm numbers between infected and control group was found on the 14th and 28th day (P < 0.001). An inverse correlation between the IgG levels and the resistance to re‐infection was also observed (r = − 0.948, P = 0.004), indicating that the resistance to reinfection is highly associated with the levels of IgG during the early phase of infection, and then with the IgM and IgE.

Occurrence of the spinal reflex due to skin pressure in sudomotor and cutaneous vasoconstrictor nerve system of humans.

The effects of skin pressure applied to one side of the waist on sudomotor and vasoconstrictor nerve activity were compared with the effects on sweating and cutaneous blood flow in humans. The sweat rate and cutaneous blood flow were measured on left and right dorsal feet. Skin sympathetic nerve activity (SSNA) was recorded by microneurography from a microelectrode inserted in left and right peroneal nerves. Skin pressure was applied in a supine position to the area over the left or right anterior superior iliac spine under warm (T(a): 30-36 degrees C) and cool (T(a): 19-23 degrees C) conditions. Sudomotor and vasoconstrictor bursts were identified for quantitative analysis. The skin pressure increased the contralateral/ipsilateral ratio of the sweat rate. It also increased the contralateral/ipsilateral ratio of the cutaneous blood flow and the contralateral/ipsilateral ratio of the sudomotor burst amplitude. However, skin pressure did not induce any significant changes in the contralateral/ipsilateral ratio of the vasoconstrictor burst amplitude. The results indicate that an asymmetrical reflex effect of skin pressure on vasoconstrictor nerve activity was absent, suggesting that, whereas the ipsilateral suppression of sweating elicited by skin pressure was mediated by the sudomotor nerve system, the ipsilateral suppression of cutaneous blood flow was not mediated by the vasoconstrictor nerve system. Thus, the occurrence of the spinal reflex due to skin pressure is not uniform between the sudomotor and the vasoconstrictor nerve systems, which represent different organizations at the level of spinal cord.

Effects of body posture on local sweating and sudomotor outflow as estimated using sweat expulsion

To estimate the effects of changes in body posture on sudomotor function, sweat rates on the forearm, chest and thigh, tympanic temperature (Tty), and skin temperatures were recorded in an upright sitting and a supine position under a hot environment of 40 degrees C Ta and 40% relative humidity for 60 min. Sweat expulsions were identified on sweat rate curves and their rates (Fsw) were calculated. Tty was higher, and its initial fall was greater, in the supine position than in the sitting position. On the forearm and the chest, the regression line relating sweat rate to mean body temperature (Tmb) had a gentler slope in the supine position, whereas on the thigh, it showed a steeper slope. The regression line relating Fsw to Tmb had a steeper slope in the supine position than in the sitting position, suggesting that the gain in the mechanisms for central integration and rhythm-generation was enhanced in the supine position. The parameter of sweat rate divided by Fsw was lower on the forearm and the chest, whereas it was higher on the thigh in the supine position than in the sitting position, suggesting that sudomotor outflow was modified at the spinal cord in association with skin pressure. It was concluded that body posture affects sudomotor functions through both brain and spinal mechanisms.