Iron chelators may help prevent photoaging.

For years, cosmetic ingredients for anti-aging treatments have attracted consumers. Skin aging is accelerated by reactive oxygen species (ROS), generated by exposure to solar ultraviolet radiation (UVR), in a process known as photoaging. Because cutaneous iron catalyses ROS generation, it is thought to play a key role in photoaging. Iron is essential to almost all forms of life. However, excess iron is potentially toxic as its catalytic activity induces the generation of ROS. Iron-catalysed ROS generation is involved in numerous pathological conditions, including cutaneous damage. When skin is directly exposed to UVR, cutaneous intracellular catalytic iron levels increase because of the release of iron from iron-binding proteins such as ferritin. Consequently, the subsequent ROS generation may overwhelm cutaneous defense systems such as the cellular iron sequestration and ROS scavenging capacity. The harmful role of excess cutaneous iron implies that there may be a potential for topical iron chelator treatments. We now consider cutaneous photodamage skin photoaging as the result of iron-catalysed ROS generation and discuss preventative strategies based on iron chelators.

Dieldrin induces apoptosis by promoting caspase-3-dependent proteolytic cleavage of protein kinase Cdelta in dopaminergic cells: relevance to oxidative stress and dopaminergic degeneration.

We previously reported that dieldrin, one of the potential environmental risk factors for development of Parkinson's disease, induces apoptosis in dopaminergic cells by generating oxidative stress. Here, we demonstrate that the caspase-3-dependent proteolytic activation of protein kinase Cdelta (PKCdelta) mediates as well as regulates the dieldrin-induced apoptotic cascade in dopaminergic cells. Exposure of PC12 cells to dieldrin (100-300 microM) results in the rapid release of cytochrome C, followed by the activation of caspase-9 and caspase-3 in a time- and dose-dependent manner. The superoxide dismutase mimetic Mn(III)tetrakis(4-benzoic acid)porphyrin chloride significantly attenuates dieldrin-induced cytochrome C release, indicating that reactive oxygen species may contribute to the activation of pro-apoptotic factors. Interestingly, dieldrin proteolytically cleaves native PKCdelta into a 41 kDa catalytic subunit and a 38 kDa regulatory subunit to activate the kinase. The dieldrin-induced proteolytic cleavage of PKCdelta and induction of kinase activity are completely inhibited by pretreatment with 50-100 microM concentrations of the caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK) and benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone (Z-DEVD-FMK), indicating that the proteolytic activation of PKCdelta is caspase-3-dependent. Additionally, Z-VAD-FMK, Z-DEVD-FMK or the PKCdelta specific inhibitor rottlerin almost completely block dieldrin-induced DNA fragmentation. Because dieldrin dramatically increases (40-80-fold) caspase-3 activity, we examined whether proteolytically activated PKCdelta amplifies caspase-3 via positive feedback activation. The PKCdelta inhibitor rottlerin (3-20 microM) dose-dependently attenuates dieldrin-induced caspase-3 activity, suggesting positive feedback activation of caspase-3 by PKCdelta. Indeed, delivery of catalytically active recombinant PKCdelta via a protein delivery system significantly activates caspase-3 in PC12 cells. Finally, overexpression of the kinase-inactive PKCdelta(K376R) mutant in rat mesencephalic dopaminergic neuronal cells attenuates dieldrin-induced caspase-3 activity and DNA fragmentation, further confirming the pro-apoptotic function of PKCdelta in dopaminergic cells. Together, we conclude that caspase-3-dependent proteolytic activation of PKCdelta is a critical event in dieldrin-induced apoptotic cell death in dopaminergic cells.

Proteolytic activation of proapoptotic kinase PKCdelta is regulated by overexpression of Bcl-2: implications for oxidative stress and environmental factors in Parkinson's disease.

We previously demonstrated that the organochlorine pesticide dieldrin, a potential chemical risk factor for development of Parkinson's disease (PD), impairs mitochondrial function and promotes apoptosis in dopaminergic PC12 cells. We further demonstrated that caspase-3-dependent proteolytic activation of a member of the novel PKC family, protein kinase Cdelta (PKCdelta), contributes to apoptotic cell death in dopaminergic cells. In the present study, we report that the proapoptotic function of PKCdelta can be regulated by overexpression of the mitochondrial anti-apoptotic protein Bcl2 in dieldrin-treated dopaminergic cells. Exposure to dieldrin (30 or 100 micro M) for 3 h produced a dose-dependent increase in caspase-3 activation and DNA fragmentation in vector-transfected PC12 cells. Overexpression of human Bcl-2 in PC12 cells completely suppressed dieldrin-induced caspase-3 activation and DNA fragmentation. Furthermore, dieldrin-induced proteolytic activation of PKCdelta was also remarkably reduced in Bcl-2-overexpressed cells. Together, these results suggest that the proapoptotic function of PKCdelta can be regulated by mitochondrial redox modulators during neurodegenerative processes.

Dieldrin-induced oxidative stress and neurochemical changes contribute to apoptopic cell death in dopaminergic cells.

We examined the acute toxicity of dieldrin, a possible environmental risk factor of Parkinson's disease, in a dopaminergic cell model, PC12 cells, to determine early cellular events underlying the pesticide-induced degenerative processes. EC(50) for 1 h dieldrin exposure was 143 microM for PC12 cells, whereas EC(50) for non-dopaminergic cells was 292-351 microM, indicating that dieldrin is more toxic to dopaminergic cells. Dieldrin also induced rapid, dose-dependent releases of dopamine and its metabolite, DOPAC, resulting in depletion of intracellular dopamine. Additionally, dieldrin exposure caused depolarization of mitochondrial membrane potential in a dose-dependent manner. Flow cytometric analysis showed generation of reactive oxygen species (ROS) within 5 min of dieldrin treatment, and significant increases in lipid peroxidation were also detected following 1 h exposure. ROS generation was remarkably inhibited in the presence of SOD. Dieldrin-induced apoptosis was significantly attenuated by both SOD and MnTBAP (SOD mimetic), suggesting that dieldrin-induced superoxide radicals serve as important signals in initiation of apoptosis. Furthermore, pretreatment with deprenyl (MAO-inhibitor) or alpha-methyl-L-p-tyrosine (TH-inhibitor) also suppressed dieldrin-induced ROS generation and DNA fragmentation. Taken together, these results suggest that rapid release of dopamine and generation of ROS are early cellular events that may account for dieldrin-induced apoptotic cell death in dopaminergic cells.

Protection against murine coxsackievirus B3 myocarditis by T cell vaccination.

T cell vaccination regulates autoimmunity by the modification of helper and suppressor T cells. The present study was performed to examine whether T cell vaccination can prevent viral myocarditis in vivo. We used coxsackievirus B3 myocarditis in mice as an animal model with the analysis of lymphokine-activated killer cell activity. Vaccination of the mice with T lymphocytes significantly prolonged survival and improved cardiac histology of murine myocarditis. The effects of T cell vaccination were most evident when T cells sensitized with the same virus were used. Vaccination of the mice with T cells from other strains of mice showed lesser protective effects. Clearance of myocardial virus was not affected by this treatment. The efficacy of T cell vaccination was confirmed in vitro by the decrease of the lymphokine-activated killer cell activity against EL-4 tumor cells and cultured myocytes. T cell vaccination of mice prolonged survival and improved myocardial lesions of animals inoculated with coxsackievirus B3.

Reduction of ultraviolet light-induced oxidative stress by amino acid-based iron chelators.

The generation of free radicals by ultraviolet (UV) light accelerates skin aging, which is known as photoaging. Cutaneous iron catalyzes the generation of free radicals. We designed novel antioxidants that suppressed the iron-catalyzed free radical generation and the ensuing UV-induced damage by mimicking the binding site of iron sequestering proteins. These antioxidants, N-(2-hydroxybenzyl)amino acids, were prepared by condensation of amino acids such as glycine and L-serine with salicylaldehyde and followed by catalytic reduction. The compounds formed a 2:1 complex to iron ion. These amino acid derivatives inhibited the iron-induced hydroxyl radical generation (the Fenton reaction). The compounds also suppressed UV-induced lipid peroxidation in murine dermal fibroblast homogenates. In addition, N-(2-hydroxybenzyl)-L-serine showed protective activity against UV-induced cytotoxicity in murine dermal fibroblasts. Desferrioxamine, a strong iron sequestering compound, was effective in inhibiting the Fenton reaction and the lipid peroxidation, but it was ineffective in protecting against UV-induced cytotoxicity. The results suggest that UV-induced oxidative stress can be reduced by these amino acid derivatives.

KMD-3213, a uroselective and long-acting alpha(1a)-adrenoceptor antagonist, tested in a novel rat model.

KMD-3213, an alpha(1a)-adrenoceptor (AR) antagonist, is under development for the treatment of urinary outlet obstruction in patients with benign prostatic hypertrophy. In the present study, we developed a rat model to investigate simply the effects of alpha(1)-AR antagonists on the intraurethral pressure (IUP) response to phenylephrine. Using this model, inhibitory effects of both i.v. and intraduodenally administered KMD-3213 on the IUP response were evaluated and compared to those of other reference compounds, including prazosin and tamsulosin. In addition, the hypotensive effects of these compounds were estimated to evaluate uroselectivity. Intravenously administered alpha(1)-AR antagonists tested, including KMD-3213, potently inhibited the IUP response in a dose-dependent manner. Although the higher doses of those compounds almost completely inhibited the IUP response, yohimbine failed to inhibit the response. When the in vivo potencies of those compounds on IUP response were correlated with their affinities for the human or animal recombinant alpha(1)-AR subtypes, alpha(1a)-AR gave the best correlation. In this model, KMD-3213 had greater uroselectivity than any other compounds examined, by both i.v. and intraduodenal routes. Moreover, 12, 18, and 24 h after the oral administration of KMD-3213, a dose-dependent inhibition of the IUP response was found, whereas the effect of tamsulosin disappeared at 18 h after the oral administration. These data indicate that KMD-3213 is a highly uroselective alpha(1)-AR antagonist with a longer duration of action. In addition, this model is useful for not only estimation of uroselectivity but also some part of the administration, distribution, metabolism, and excretion of many compounds to discover uroselective compounds.

Antioxidants modulate acute solar ultraviolet radiation-induced NF-kappa-B activation in a human keratinocyte cell line.

Exposure of the human skin to ultraviolet radiation (UVR) leads to depletion of cutaneous antioxidants, regulation of gene expression and ultimately to the development of skin diseases. Although exogenous supplementation of antioxidants prevents UVR-induced photooxidative damage, their effects on components of cell signalling pathways leading to gene expression has not been clearly established. In the present study, the effects of the antioxidants alpha-lipoic acid, N-acetyl-L-cysteine (NAC) and the flavonoid extract silymarin were investigated for their ability to modulate the activation of the transcription factors nuclear factor kappa B (NF-kappaB) and activator protein-1 (AP-1) in HaCaT keratinocytes after exposure to a solar UV simulator. The activation of NF-kappaB and AP-1 showed a similar temporal pattern: activation was detected 2 h after UV exposure and maintained for up to 8 h. To determine the capacity of activated NF-kappaB to stimulate transcription, NF-kappaB-dependent gene expression was measured using a reporter gene assay. The effects of the antioxidants on NF-kappaB and AP-1 activation were evaluated 3 h after exposure. While a high concentration of NAC could achieve a complete inhibition, low concentrations of alpha-lipoic acid and silymarin were shown to significantly inhibit NF-kappaB activation. In contrast, AP-1 activation was only partially inhibited by NAC, and not at all by alpha-lipoic acid or silymarin. These results indicate that antioxidants such as alpha-lipoic acid and silymarin can efficiently modulate the cellular response to UVR through their selective action on NF-kappaB activation.

Reversed circadian blood pressure rhythm is associated with occurrences of both fatal and nonfatal vascular events in NIDDM subjects.

To assess the significance of reversed circadian blood pressure (BP) rhythms as a predictive factor of vascular events in NIDDM, vital status after an average 4-year follow-up was determined in 325 NIDDM subjects in whom the circadian BP profile had been monitored between 1988 and 1996. Circadian BP rhythm was analyzed by the COSINOR (a compound word for cosine and vector) method, as previously described. After exclusion of 37 dropped-out subjects, 288 were recruited to the further analysis, of which 201 had a normal circadian BP rhythm (group N) and the remaining 87 had a reversed one (group R). There was no difference in sex, HbA1c, the prevalence of smokers, serum lipids, or serum electrolytes between groups N and R at baseline, whereas age, the prevalence of hypertension, serum creatinine, and diabetic complications were more pronounced in group R than in group N. During the follow-up period (which averaged 52 months in group N and 36 months in group R), fatal and nonfatal vascular (cerebrovascular, cardiovascular, peripheral vascular arteries, and retinal artery) events occurred in 20 subjects in group N and 56 in group R. Unadjusted survival times and event-free times were estimated by the Kaplan-Meier product-limit method, and there was a significant difference in both unadjusted survival and event-free survival rates between groups N and R (P < 0.001 each; log-rank test). The Cox proportional-hazards model adjusted for age, sex, circadian BP pattern, duration of diabetes, therapy for diabetes, various diabetic complications, and hypertension demonstrated that circadian BP pattern and age exhibited significant, high adjusted relative risks for fatal events, and that diabetic nephropathy, postural hypotension, and hypertension as well as circadian BP pattern exhibited significant, high adjusted relative risks with respect to the occurrence of various nonfatal vascular events. These results suggest that reversed circadian BP rhythm is associated with occurrences of both fatal and nonfatal vascular events in NIDDM subjects.

Rapidly progressive glomerulonephritis concomitant with diabetic nephropathy.

We describe a rare case of a rapidly progressive glomerulonephritis (RPGN) superimposed on diabetic nephropathy. A 68-year-old woman with non-insulin-dependent diabetes mellitus (NIDDM) complicated with diabetic triopathy demonstrated a rapid deterioration of renal function. Her urinary sediment contained many red blood cell (RBC) cells and casts, suggesting an additional renal disease accompanying diabetic nephropathy. Renal biopsy revealed crescent formation in many glomeruli characteristic of the pauci-immune type of RPGN. Steroid pulse therapy transiently halted the deterioration in renal function, but the patient died of pneumonia complicated with methicillin-resistant staphylococcus aureus (MRSA) infection. The unusual findings in diabetic nephropathy indicated the coexistence of primary glomerulonephritis and diabetic glomerulosclerosis in this case.

Suppression of iron catalyzed free radical generation by iron tyrosinate protein models.

Novel antioxidants, N-(2-Hydroxy-1-naphthal)amino acids which mimic iron proteins, have been intensively studied for suppressive effect on iron-catalyzed free radical generation. These compounds exhibit inhibition of the Fenton reaction in electron spin resonance assessment. In addition, it is shown that the compounds inhibit iron-induced peroxidation by thiobarbituric acid test. Since these antioxidants form stable complexes with Fe3+, this antioxidative activity is expected to be derived from sequestration of catalytic iron.

[An experiment on drinking using breath alcohol monitor (Alcomed 3010) by an electrochemical method].

A drinking experiment was performed to evaluate the efficiency of a breath alcohol monitor, Alcomed 3010. The ethanol concentrations in blood and breath were determined by gas chromatography, and in particular the breath ethanol concentration was determined with the breath alcohol monitor and by gas chromatography. The results obtained by two methods were compared. Based on the blood and breath ethanol concentrations, the following conclusions were drawn reading the breath alcohol monitor. The monitor has practical merit for determination of the breath ethanol level. It is small, usable anywhere, with little error in determination. In measuring principle, tobacco and acetone did not affected levels with the meter, but methanol, n-propanol and n-butanol affected determinations with the alcohol monitor. The breath (AM)/blood (GC) ethanol ratio was 1:2555. Comparison of the values determined with the alcohol monitor and gas chromatography yielded the equation: y = 0.998 x +/- 0.012 (r = 0.994). When determinations were made on the pure ethanol gas by the meter and gas chromatograph, the equation was: y = 0.974 x +/- 0.021 (r = 0.994). It may be said therefore that the alcohol monitor is both practically and functionally excellent.

Simultaneous measurement of alcohols and hydrogen cyanide in biological specimens using headspace gas chromatography.

We attempted to analyze biological specimens simultaneously for alcohols and hydrogen cyanide. A headspace gas chromatographic method with thick film wide bore column (PEG 20M) for the simultaneous determinations of methanol, ethanol, n-propanol and hydrogen cyanide in blood has been developed. This method was applied for the determinations of methanol, ethanol and hydrogen cyanide in a forensic autopsy case and animal experiments.

[A case of amylase producing lung cancer].

The patient was a 72-year-old man, who was admitted to our hospital because of cough. Chest X-rays showed a mass shadow in the right lower lung field. Amylase activities in serum and urine were extremely high. Amylase isozyme pattern identified salivary type amylase. Cytological examination of the sputum suggested adenocarcinoma. Amylase activities in serum and urine gradually decreased with the administration of chemotherapy. Afterwards, pleural effusion increased, and the amylase activity in pleural fluid was also extremely high. Pleural fluid also showed adenocarcinoma. Enzyme-labeled antibody method (PAP) on this specimen from pleural fluid proved that tumor cells were producing amylase ectopically.

[A case of partial absence of the left pericardium].

A rare case of partial absence of the left pericardium was reported. The patient was a seventy-nine year old male and the cause of death was a gastric cancer. The pericardial absence was oval in shape, in egg-size with smooth margin and was located between the superior portion of the left pericardium and the pleural cavity. The pulmonary artery, left auricle and superior part of the left ventricle were visible through the absence. The left phrenic nerve descended along the anterior free margin of the absence.

Papillary adenoma of the lung. Histological and ultrastructural findings in two cases.

Two cases of papillary adenoma of the lung are presented along with results of histological and ultrastructural examinations. The tumors were encountered in two asymptomatic patients in a mass-survey chest X-ray examination. The chest X-ray films showed the tumors as well demarcated small lesions. Histologically, both tumors arose in the bronchioles and consisted of cuboidal cells resembling type II pneumocytes showing papillary growth with accompanying edematous connective tissue. Several tumor cells each possessed a large eosinophilic intranuclear inclusion. In case 1, ciliated cells and Clara-like cells were also present in the tumor. Ultrastructurally, most of the tumor cells had various numbers of lamellar bodies in their cytoplasm, indicative of type II pneumocytes, and some of case 1 showed features of Clara cells and ciliated cells. The intranuclear inclusions appeared as aggregates of tubular structures or had lamellar body-like features. These findings are identical to those of papillary adenoma arising from the bronchiole.

[Continuous subcutaneous infusion of GnRH agonist: effective dosage in the treatment of endometriosis and its influence on the ovarian response to human menopausal gonadotropin].

This study was designed to compare the clinical and hormonal efficacy of the treatment for endometriosis using continuous infusion of three different doses of GnRH agonist (A). In addition, we examined the ovarian responsiveness to human menopausal gonadotropin (hMG) administration during GnRH-A treatment. Thirteen endometriosis patients were divided into 3 groups and given different doses. GnRH-A (Buserelin) was infused continuously through the subcutaneous route at rates of 200 micrograms (Group I; n = 5), 100 micrograms (Group II, n = 4) and 10 micrograms (Group III; n = 4) per day for 24 weeks. After the start of treatment, serum estradiol (E2) was suppressed to the menopausal range within 2 weeks and thereafter maintained this range until 24 weeks in each group. The LH and FSH response to a GnRH Challenge test was completely abolished within 2 weeks in 3 groups. Although serum FSH decreased to below the pretreatment value within a week, the FSH level was significantly lower in groups I and II than in group III until 8 weeks. No difference in the LH level during the treatment was seen among the 3 groups. After completion of the 24 weeks' treatment, FSH increased rapidly, and ovulation returned within 4 to 6 weeks in each group. Pregnancy was achieved in two patients in group I, one patient in group II and one patient in group III during cycles 2 and 5. Serum E2 increased to 200-300 pg/ml in 3 out of 7 patients treated with hMG during GnRH-A infusion, whereas no increase in E2 was seen in the remaining 4 patients.(ABSTRACT TRUNCATED AT 250 WORDS)