Analysis of sebum lipid composition and the development of acneiform rash before and after administration of egfr inhibitor.

Treatment with an epidermal growth factor receptor inhibitor (egfri) in patients having non-small-cell lung cancer can cause frequent and diverse skin toxicities, an acneiform rash being one of the commonest. Although the exact pathophysiology of this rash and its development mechanisms remain unknown, investigators have noted that egfri-induced skin toxicity might be partly associated with sebaceous gland function. Sebum is composed mainly of the lipids squalene (sq), wax ester (we), triglyceride, free fatty acid, and cholesterol, which are secreted mostly from the sebaceous glands and by keratinocytes. We therefore investigated the lipid composition of sebum before and after administration of egfri and whether sebum composition was associated with the development of acneiform rash. To investigate any associated changes in sebum gland activity, we focused especially on alterations in the amounts of sq and we, which are secreted solely from the sebaceous glands. In contrast to our expectations, we observed no substantial changes in the lipid composition of sebum before and after administration of egfri. Composition varies with the individual; however, the proportion of sq and we derived from the sebaceous glands was significantly lower in regions that did not develop acneiform rash than in regions that did. Our results suggest that development of an acneiform rash after administration of egfri could be related to sebaceous gland activity. Measurement of the lipid composition of sebum before therapy with egfri might predict which patients will be prone to acneiform rash.

Topical application of PPARα (but not ß/δ or γ) suppresses atopic dermatitis in NC/Nga mice.

BACKGROUND: Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors, which regulate not only adipogenesis and proliferation/differentiation but also the immune response of cells. Because topical application of the activators of some PPAR isoforms improved clinical symptoms in patients with atopic dermatitis (AD), we investigated the role of PPAR activators using a murine AD model in NC/Nga mice; to the best of our knowledge, this has not been previously reported. METHODS: Activators of three PPAR isoforms (α, ß/δ, γ) were topically applied on inflamed skin in a murine AD model that was developed by repeated topical application of mite antigen in NC/Nga mice. The efficacy of each topical PPAR activator was evaluated immunologically and serologically. RESULTS: Topical application of the PPARα activator, but not of the activators of PPARß/δ or PPARγ, improved clinical dermatitis, reduced inflammatory cell infiltration in the dermis, and alleviated the elevation of serum IgE levels. In addition, PPARα expression was downregulated in the epidermis in our murine AD model, as is seen in patients with AD. CONCLUSIONS: Topical application of PPARα activator could be a potent therapeutic agent for patients with AD and could take the place of topical steroid treatments.

Expression of milk fat globule epidermal growth factor-VIII may be an indicator of poor prognosis in malignant melanoma.

BACKGROUND: Milk fat globule epidermal growth factor-VIII (MFG-E8) is a secreted protein that binds phosphatidylserine and promotes apoptotic cell ingestion by phagocytes, mediating the immune tolerance and maintenance of homeostasis. A recent study has shown that MFG-E8 expression in human melanoma is increased with tumour progression; however, the effect of its expression on patient survival has not yet been clarified. OBJECTIVE: To analyse MFG-E8 expression in melanoma, and to determine whether it can serve as a marker for diagnosis, tumour progression and/or prognosis. METHODS: MFG-E8 expression was examined by immunohistochemistry in 60 primary melanomas, 22 metastatic lesions and 30 benign naevi. The following clinicopathological variables were evaluated: age, gender, histological type, tumour site, Breslow thickness, Clark's level, the presence or absence of ulceration and tumour-infiltrating lymphocytes, and survival periods. Statistical analyses were performed to assess associations and melanoma-specific survival. RESULTS: MFG-E8 expression was significantly higher in primary and metastatic melanoma than in naevus. Furthermore, it increased according to tumour progression and metastasis. Patients with MFG-E8 expression in primary tumours had significantly shorter survival periods than those without MFG-E8 expression. Univariate and multivariate analyses revealed that MFG-E8 expression was a statistically significant and independent prognostic factor. CONCLUSION: MFG-E8 expression may serve as a tumour progression marker and can predict an unfavourable prognosis in patients with melanoma.

Stromal expression of cathepsin K in squamous cell carcinoma.

BACKGROUND: Cathepsin K (CTSK), a cysteine protease with strong collagenolytic and elastolytic properties involved in extracellular matrix turnover, may be produced by neoplastic cells as well as stromal macrophages and fibroblasts. Its expression is suggested as associated with increased invasive and metastatic potential. OBJECTIVES: The aim of this study is to examine stromal expression of cathepsin K in skin tumors. METHODS: A series of 13 normal skin and 109 skin tumours, including 51 benign and 58 malignant epidermal tumours were tested for CTSK and Ki-67 expression by immunohistochemical analysis. RESULTS: Stromal CTSK expression and the tumoral Ki-67 labelling index were significantly higher in invasive squamous cell carcinoma (SCC) than in other epidermal tumours. CONCLUSION: Cathepsin K-positive stromal fibroblasts may play a crucial role in SCC progression by promoting extracellular matrix degradation, thereby facilitating SCC growth and invasion into surrounding tissue and vasculature. CTSK inhibitors may be a potential novel therapeutic option to decrease SCC progression.

Heteroclitic immunization induces tumor immunity.

In tumor transplantation models in mice, cytotoxic T lymphocytes (CTLs) are typically the primary effector cells. CTLs recognize major histocompatibility complex (MHC) class I-associated peptides expressed by tumors, leading to tumor rejection. Peptides presented by cancer cells can originate from viral proteins, normal self-proteins regulated during differentiation, or altered proteins derived from genetic alterations. However, many tumor peptides recognized by CTLs are poor immunogens, unable to induce activation and differentiation of effector CTLs. We used MHC binding motifs and the knowledge of class I:peptide:TCR structure to design heteroclitic CTL vaccines that exploit the expression of poorly immunogenic tumor peptides. The in vivo potency of this approach was demonstrated using viral and self-(differentiation) antigens as models. First, a synthetic variant of a viral antigen was expressed as a tumor antigen, and heteroclitic immunization with peptides and DNA was used to protect against tumor challenge and elicit regression of 3-d tumors. Second, a peptide from a relevant self-antigen of the tyrosinase family expressed by melanoma cells was used to design a heteroclitic peptide vaccine that successfully induced tumor protection. These results establish the in vivo applicability of heteroclitic immunization against tumors, including immunity to poorly immunogenic self-proteins.

Distribution of kinetochore (centromere) antigen in mammalian cell nuclei.

Antigens associated with mammalian centromeres were localized at the high and electron microscopic levels using the peroxidase-labeled antibody method. The antibody used was of a type naturally occurring in the sera of patients with scleroderma. At the light microscopic level, it reacts specifically with the centromere regions of chromosomes in a variety of mammalian species and strains in discrete foci in interphase nuclei. We find that the number of foci approximates the number of chromosomes present in the various cell types. At the ultrastructural level, the antigenic foci are confirmed to lie in the kinetochore regions of each chromosome. In interphase nuclei, the antigenic foci were usually associated either with the inner surfaces of the nuclear envelope or with the nucleoli. These observations indicate that the centromere regions of the chromosomes in interphase are not randomly distributed within the nucleus but are usually fixed either to the inner surface of the nuclear envelope or to nucleoli.

Immunohistochemical analysis of the mammalian target of rapamycin signalling pathway in extramammary Paget's disease.

BACKGROUND: We have previously observed that persistent activation of the serine/threonine kinase, protein kinase B (AKT) is a frequent event in extramammary Paget's disease (EMPD). AKT promotes cell proliferation by its ability to coordinate mitogenic signalling with energy- and nutrient-sensing pathways that control protein synthesis through the atypical serine/threonine kinase, mammalian target of rapamycin (mTOR). CDK2, a member of the serine/threonine kinase family of cyclin-dependent kinases, is a key regulator of G(1)-S cell cycle progression, and has recently been shown to be one of the targets of AKT. The AKT-mTOR-p70 ribosomal protein S6 kinase (p70S6K) pathway has been described in some human malignancies, but not in EMPD. OBJECTIVE: To investigate the immunohistochemical staining of the AKT-mTOR-p70S6K pathway in EMPD and to evaluate the relationships among the components. METHODS: Samples of primary EMPD tissue were subjected to immunohistological staining with phosphorylated (p)-AKT, p-mTOR, p-4E-binding protein 1 (p-4EBP1), p-p70S6K/S6K1, p-ribosomal protein S6 (p-S6) and CDK2. Ten normal skin samples served as a control. RESULTS: Of the 32 EMPD tissue samples, 29, 27, 26, 29, 26 and 32 samples were positive for p-AKT, p-mTOR, p-4EBP1, p-p70S6K/S6K1, p-S6 and CDK2 staining, respectively. All these cell signalling molecules showed higher positivity in invasive EMPD than in EMPD in situ. There were significant correlations between p-AKT, p-mTOR, p-4EBP1, p-p70S6K/S6K1 and p-S6 and CDK2. CONCLUSIONS: The activation of the AKT-mTOR-p70S6K pathway may play an important role in the pathogenesis of EMPD. The high expression of the components of the pathway was highly correlated with CDK2 expression, suggesting that the AKT/mTOR pathway may induce the malignant transition through CDK2 in EMPD. The AKT-mTOR-p70S6K pathway might be a potential therapeutic target in EMPD.

Activation of the mammalian target of rapamycin signalling pathway in epidermal tumours and its correlation with cyclin-dependent kinase 2.

BACKGROUND: The enzyme mammalian target of rapamycin (mTOR) integrates many different cellular signals to control cell growth and proliferation, protein synthesis and breakdown, and other processes. Dysregulation of mTOR is implicated in a range of human diseases, including cancers and cardiovascular disorders. To date, there has been no report on the expression of protein kinase B (AKT)/mTOR cell signalling in epidermal tumours. OBJECTIVES: This study was designed to investigate the activation of the mTOR signalling pathway in epidermal tumours and to correlate this with cyclin-dependent kinase 2 (CDK2) expression. METHODS: Immunohistological staining was performed with phosphorylated (p-) AKT, p-mTOR, p-4E-binding protein 1 (p-4EBP1), p-ribosomal protein S6 (p-S6), p-p70 ribosomal protein S6 kinase 1 (p-p70S6K1) and CDK2 in 15 cases each of seborrhoeic keratosis, actinic keratosis, keratoacanthoma and Bowen's disease (BD), and 25 cases of squamous cell carcinoma (SCC). Fifteen normal skin (NS) samples served as control. RESULTS: Among 85 tumours, 40 (47%) were positive for p-AKT, 31 (36%) for p-mTOR, 44 (52%) for p-4EBP1, 38 (45%) for p-S6, and 39 (46%) for p-p70S6K1. CDK2 immunostaining was positive in all cases of SCC and BD, and in 67% of benign tumours. All of these markers were stained much more frequently in malignant tumours than in benign tumours or NS. p-AKT, p-mTOR, p-4EBP1, p-p70S6K1 and p-S6 each showed high correlation with CDK2. CONCLUSIONS: Constitutive activation of the AKT/mTOR pathway was frequent in epidermal tumours, especially in malignant tumours. Activation was highly correlated with CDK2 expression, suggesting that the AKT/mTOR pathway may induce the malignant transition through CDK2 in epidermal tumours.

Increased expression of an epidermal stem cell marker, cytokeratin 19, in cutaneous squamous cell carcinoma.

BACKGROUND: Cytokeratin 19 (CK19) has been considered to be a putative marker for epidermal stem cells in the hair follicle bulge. Cumulative reports have shown that epidermal stem cells play an important role in skin carcinogenesis. However, to date there has been no report on the clinical alteration of the stem cells in squamous cell carcinoma (SCC). OBJECTIVES: To investigate alteration of the stem cells and proliferating cells and to assess their relationship and potential contribution to SCC. METHODS: Thirty paraffin-embedded neoplastic skin lesions, consisting of 10 cases each of actinic keratosis (AK), Bowen disease (BD) and SCC, were examined immunohistologically for CK19 and Ki-67. RESULTS: Positive reactivity for CK19 was seen in 30% of AK, 50% of BD and 80% of SCC lesions. There was significantly higher expression levels of CK19 in SCC than in AK and BD (P < 0.05). In addition, BD lesions harboured a significantly higher number of CK19-positive cells than did AK lesions (P < 0.05). There were significant differences in Ki-67 labelling indices between AK and BD and between AK and SCC (P < 0.001), but not between BD and SCC (P > 0.05). Furthermore, a serial section comparison study showed that there was a minor population of cells co-expressing CK19 and Ki-67 in a subset of the tumour cells of SCC samples. The percentage of CK19+ cells significantly correlated with that of Ki67+ cells in all examined neoplastic skin lesions. CONCLUSIONS: These results suggest that CK19 expression may be associated with the retention of stem cell characteristics or a state that is uncommitted to terminal squamous differentiation.

Differential expression of two new members of the p53 family, p63 and p73, in extramammary Paget's disease.

BACKGROUND: The proteins p53, p63 and p73 are known to be overexpressed and to play important roles in the pathogenesis of many tumours, but the expression of p63 and p73 has not previously been investigated in extramammary Paget's disease (EMPD). AIM: To investigate the potential contribution of p53, p63 and p73 in the pathogenesis of EMPD. METHODS: In total, 35 paraffin wax-embedded tissue samples from patients with EMPD were examined using immunohistochemical staining for p53, p63 and p73. RESULTS: All of the 35 EMPD specimens, including all 6 invasive EMPD and 2 metastatic lymph-node specimens, showed nuclear overexpression of both p53 and p73. The expression levels (percentage of positive cells) of p53 and p73 (90.66 +/- 12.53% and 80.20 +/- 13.07%) in EMPD were significantly higher than those of normal skin. There was a significant correlation between the expression levels of p53 and p73 in EMPD. In 29 of 35 EMPD specimens, there was no nuclear expression of p63, and weak or moderate staining was found in only 6 specimens. The expression level of p63 in EMPD was significantly less than that in normal skin. CONCLUSIONS: Our study shows that the concordant overexpression of p53 and p73 and the decreased expression of p63 may play a pivotal role in the pathogenesis of EMPD. The decreased expression of p63 may play a more important role in the pathogenesis of EMPD than the overexpression of p53 and p73.

Tumor immunity and autoimmunity induced by immunization with homologous DNA.

The immune system can recognize self antigens expressed by cancer cells. Differentiation antigens are prototypes of these self antigens, being expressed by cancer cells and their normal cell counterparts. The tyrosinase family proteins are well characterized differentiation antigens recognized by antibodies and T cells of patients with melanoma. However, immune tolerance may prevent immunity directed against these antigens. Immunity to the brown locus protein, gp75/ tyrosinase-related protein-1, was investigated in a syngeneic mouse model. C57BL/6 mice, which are tolerant to gp75, generated autoantibodies against gp75 after immunization with DNA encoding human gp75 but not syngeneic mouse gp75. Priming with human gp75 DNA broke tolerance to mouse gp75. Immunity against mouse gp75 provided significant tumor protection. Manifestations of autoimmunity were observed, characterized by coat depigmentation. Rejection of tumor challenge required CD4(+) and NK1.1(+) cells and Fc receptor gamma-chain, but depigmentation did not require these components. Thus, immunization with homologous DNA broke tolerance against mouse gp75, possibly by providing help from CD4(+) T cells. Mechanisms required for tumor protection were not necessary for autoimmunity, demonstrating that tumor immunity can be uncoupled from autoimmune manifestations.

Micelle formation of sodium cholate and solubilization into the micelle.

The micellization of sodium cholate (NaC) was studied at 298.2 K by aqueous solubility at different pH values. Using a stepwise association model of cholate anions without the sodium counterion, the aggregation number (n) of the cholate micelle was evaluated and found to increase with the total concentration, indicating that the mass action model worked quite well. The n value at 60 mM was found equal to 16. The membrane potential measurement of sodium ion with a cation exchange membrane was made in order to confirm the low counterion binding to micelle. The solubilization of alkylbenzenes (benzene, toluene, ethylbenzene, n-propylbenzene, n-butylbenzene, n-pentylbenzene, n-hexylbenzene) and polycyclic aromatic compounds (naphthalene, anthracene, pyrene) into the aqueous micellar solution of sodium cholate was carried out. Solubilizate concentrations at equilibrium were determined spectrophotometrically at 298.2 K. The first stepwise association constants (K1) between solubilizate monomer and vacant micelle were evaluated from the equilibrium concentrations and found to increase with increasing hydrophobicity of the solubilizate molecules. From the Gibbs energy change for solubilization at the different mean aggregation numbers and from molecular structure of the solubilizates, the function of sodium cholate micelle for solubilization was discussed and was compared with data from conventional aliphatic micelles.

Ultraviolet-B irradiation alters cytokine production by immune lymphocytes in herpes simplex virus-infected mice.

Previous studies from our laboratories have shown that UV-B irradiation at the site of an intradermal infection of herpes simplex virus (HSV) resulted in a higher incidence of zosteriform lesions and suppressed cellular immune responses to HSV in mice. In order to determine whether the production of T-cell-derived cytokine (IFN-gamma, IL-2 and IL-4) by immune cells from irradiated mice is also suppressed, we examined the production of cytokines by lymph node cells and spleen cells taken from UV-B irradiated, HSV type 1 (HSV-1)-infected mice. UV-B irradiation (120 mJ/cm2) prior to HSV-1 infection was found to markedly suppress IFN-gamma production compared to that of the non-irradiated control. IL-4 production was enhanced compared to IL-2 in the UV-B irradiated mice. These results suggest that alteration(s) in the cytokine production profile may therefore be involved in the development of severe skin lesions caused by HSV infection in UV-B irradiated mice.

Detection of beta-globin mRNA precursor in heterogeneous nuclear ribonucleoprotein associated with U1-RNP by using anti-RNP antibody.

Heterogeneous nuclear RNA-ribonucleoprotein (hnRNP) fractions were isolated from Friend erythroleukemia cells and separated by 15-45% sucrose gradient centrifugation. The distribution of small nuclear RNAs (snRNAs) in hnRNP fractions indicated that the snRNAs are associated with hnRNP particles. HnRNP fractions were incubated with normal IgG or anti-U1 RNP IgG, and the resulting immunocomplexes were isolated by binding to a protein A-Sepharose column. HnRNP was found in bound fractions only when anti-U1 RNP IgG was used. By Northern hybridization of RNA extracted from the immunocomplexes with a beta-globin genomic DNA probe, 15S beta-globin mRNA precursors and 10S mature mRNA were detected. These findings suggest the existence of a complex of U1 RNP particles and hnRNP particles containing beta-globin pre-mRNA.

Augmentation of in vitro cytolytic activity of LAK cells with heated ATL-derived cell lines.

Three adult T-cell leukemia/lymphoma-derived cell lines, MT-2, MJ, and HUT102, were investigated to determine how they responded to hyperthermia, lymphokine-activated killer (LAK) cells, or a combination of both in vitro. All three cell lines showed a similar sensitivity to LAK cells, but revealed varying degrees of sensitivity to hyperthermia (MT-2 < MJ < HUT102) by 51Cr release assay. Hyperthermia did not cause immediate cell death as determined by the trypan blue exclusion test, but did cause substantial decreases in the numbers of heated cells within 2 days. The density of the cells began to increase thereafter, which was consistent with the results of the experiments labeling the cells with 3H-TdR after hyperthermia. When the cells were heated at 39-43 degrees C for 1-3 hr and then interacted with various LAK cell/ATL cell (E/T) ratios at 37 degrees C for 4 hr, total cytolysis of the cells increased in a synergistic and/or additive manner over that of the cells without hyperthermia. Prolonged incubation of the cells at high temperature did not necessarily cause a large increase in the interaction of LAK cells after hyperthermia. This augmentation of cytolysis by LAK cells after hyperthermia was not seen in normal peripheral lymphocytes. These results suggest that the combination therapy of hyperthermia and LAK cells may be more specific, useful, and effective for treating malignant lymphoma.

[Quality of life, subjective health status and health and life satisfaction in rheumatoid arthritis].

A Japanese version of Arthritis Impact Measurement Scales (AIMS) was developed after the original AIMS Version 2 and utilized for Quality of Life (QOL) measurement in 691 patients with Rheumatoid Arthritis (RA). Various medical (physical and laboratory) examinations, which are widely used in the clinical settings for the assessment of RA activity and severity, were also performed by physicians. Interrelationships between QOL, patient subjective health status, and health and life satisfaction were analyzed with the following results: 1: The effect of QOL impairment by RA upon patients' subjective health rating and health satisfaction were not constant over the range of severity of disease status. Pain was found to lower overall subjective health and health satisfaction regardless of RA class. On the other hand, while the deterioration of mobility aspects of QOL had negative effects upon patients' subjective health status and satisfaction among less-disabled RA patients, any of physical aspects of QOL, including the degree of mobility impairment, showed no significant association with patients' subjective health status and satisfaction in the more disabled. 2: Psychological aspects of QOL (mood and tension) had significant associations with patients' subjective health status and satisfaction. In the less severe group, mood impairment had a significant effect on subjective health and satisfaction, while in the more severe group tension showed a significant association. It was indicated that management of psychological aspects of QOL is important in RA patients to improve and advance their subjective health status and satisfaction. 3: Although social aspects of QOL, i.e. social support, social life and job status, showed no significant relationship to subjective health rating and health satisfaction, those with less disease severity who lacked social support and who had a jobless state were likely to have lower disease acceptance and life satisfaction, while those with more severe disease who had less social interaction manifested lower life satisfaction. These results suggested that social aspects of QOL, while not directly associated with subjective health rating, could be important factors affecting disease acceptance and life satisfaction.

A Case of Sparganosis mansoni in the Thigh: Serological Validation of Cure Following Surgery.

Cases of Sparganum mansoni, caused by the plerocercoid larva of the tapeworm S. mansoni, occur throughout the world, particularly in Asian, Middle Eastern, and European countries. However, cases of infection with this parasite are rarely seen in Japan. Here, we present a case of a 61-year-old woman with a solitary subcutaneous nodule in left inner aspect of the thigh, from which a long, slender, whitish worm was surgically removed. The parasite was histopathologically identified as S. mansoni. Serological testing confirmed cure of the infection after surgical removal of the parasite. The authors advocate immunoserological examination in case of S. mansoni.