Positive correlation between PEDF expression levels and macrophage density in the human prostate.

BACKGROUND: In this study, we investigated the capacity of pigment epithelium-derived factor (PEDF) to modulate the recruitment and the differentiation of monocytes/macrophages both in vitro and in human prostate. METHODS: Using Boyden chambers, we assessed PEDF effect on the migration of monocytes and chemically activated RAW 264.7 macrophages. Normal, prostatitis, and prostate cancer specimens were retrospectively selected and examined by immunohistochemistry for PEDF expression and infiltration of immune CD68 + macrophagic cells. PEDF expression and macrophage density were then correlated with each other and clinicopathological parameters. M1 and M2 differentiation markers were quantified by qRT-PCR, Western blotting, and ELISA. RESULTS: In chemotaxis, PEDF induced the migration of monocytes/macrophages. In immunohistochemistry, macrophages were markedly increased in prostatitis and malignant compared to normal tissues. PEDF was expressed at variable levels in the stroma and epithelium. PEDF mRNA was down-regulated in both prostate cancer and prostatitis compared to normal tissues. In correlation studies, macrophage density and PEDF expression were respectively positively and negatively associated with prostate size. Most importantly, PEDF expression positively correlated with macrophage density. Finally, PEDF stimulated the expression of iNOS, IL12, and TNFα; and inhibited IL10 and arginase 1 in mouse and human macrophages confirming a M1-type differentiation. CONCLUSIONS: Our data demonstrate that PEDF acts directly on monocytes/macrophages by inducing their migration and differentiation into M1-type cells. These findings suggest a possible role of macrophages in PEDF anti-tumor properties and may support further development of PEDF-based anti-cancer therapy.

Cutaneous phaeohyphomycosis caused by Paraconiothyrium cyclothyrioides.

Paraconiothyrium cyclothyrioides is a recently described coelomycetous fungal species. We present a case in a renal transplant patient with chronic skin lesions of the lower extremities caused by P. cyclothyrioides. Treatment with posaconazole led to complete resolution of the lesions. P. cyclothyrioides should be considered an opportunistic human pathogen in immunocompromised patients.

Green tea polyphenols and 1-α-OH-vitamin D3 attenuate chronic inflammation-induced myocardial fibrosis in female rats.

Studies have suggested that 1-α-OH-vitamin D3 and green tea polyphenols (GTPs) are promising dietary supplements for mitigating chronic inflammation-induced fibrosis of vessels because of their anti-inflammatory properties. This study evaluated (1) the impact of 1-α-OH-vitamin D3 on myocardial fibrosis in female rats with chronic inflammation and (2) if 1-α-OH-vitamin D3 and GTPs have an additive or synergistic effect to attenuate myocardial fibrosis in these female rats. A 3-month study of a 2 (no 1-α-OH-vitamin D3 vs. 0.05 µg/kg 1-α-OH-vitamin D3, five times per week) ×2 (no GTPs vs. 0.5% GTPs in drinking water) factorial design in lipopolysaccharide (LPS)-administered female rats was performed. Additionally, a group receiving placebo administration was used to compare with a group receiving LPS administration only to evaluate the effect of LPS. Masson's Trichrome staining evaluated myocardial fibrosis in coronary vessels and surrounding myocardium. Spleen cyclooxygenase-2 mRNA expression was determined by real-time polymerase chain reaction. Total lipid profiles were also determined. Whole blood was used for differential cell counts. Data were analyzed by two-way analysis of variance followed by mean separation procedures. At 3 months LPS administration induced myocardial fibrosis in vessels and surrounding myocardium, spleen cyclooxygenase-2 mRNA expression, and elevated leukocyte counts, whereas both 1-α-OH-vitamin D3 administration and GTPs supplementation significantly attenuated these pro-inflammatory events. The inhibitory effects of 1-α-OH-vitamin D3 and GTPs seem to be an individual effect, instead of an additive or synergistic effect. 1-α-OH-vitamin D3 and GTPs lowered red blood cell counts, hematocrit, and hemoglobin. Neither 1-α-OH-vitamin D3 nor GTPs affected lipid profiles. In summary, both 1-α-OH-vitamin D3 administration and GTPs supplementation mitigate myocardial fibrosis through suppression of a chronic inflammation innate immune response.

CRM1-dependent p53 nuclear accumulation in lung lesions of a bitransgenic mouse lung tumor model.

The p53 tumor suppressor gene plays an essential role in tumorigenesis, and the chromosomal region maintenance 1 (CRM1) has been suggested to export p53 protein from the nucleus to the cytoplasm. The objectives of the present study were to evaluate p53 expression and subcellular localization as well as CRM1 expression using immunohistochemistry in our established bitransgenic mouse lung tumor model. In this model, expression of the mutant human Ki-rasG12C allele was regulated in a doxycycline (DOX)-inducible, lung-specific manner. Following treatment with curcumin, we found that although overall p53 expression levels were not significantly changed among the three groups, lung lesions in mice treated with DOX alone had the highest proportion of N>C (nucleus predominant) p53 staining (46±7%), followed by lung lesions in mice co-treated with DOX and curcumin (31±12%) and controls (17±4%). CRM1 expression was dramatically inhibited in lung lesions in mice treated with DOX (0±0) as compared to controls (90±17, P=0.001), and could be partially reversed after curcumin treatment (47±21, P=0.028, DOX vs. DOX+curcumin). Collectively, the results from this study demonstrated that p53 accumulated in the nucleus in lung lesions in mice expressing the mutant Ki-rasG12C transgene as a result of down-regulation of CRM1. Furthermore, these alterations could be partially reversed by curcumin treatment. p53 subcellular localization resulting from CRM1 alterations may play an important role in lung tumorigenesis.

Green tea polyphenols avert chronic inflammation-induced myocardial fibrosis of female rats.

OBJECTIVE: Green tea proposes anti-inflammatory properties which may attenuate chronic inflammation-induced fibrosis of vessels. This study evaluated whether green tea polyphenols (GTP) can avert fibrosis or vascular disruption along with mechanisms in rats with chronic inflammation. TREATMENTS: Forty 3-month-old female rats were assigned to a 2 (placebo vs. lipopolysaccharide, administration) × 2 (no GTP vs. 0.5% GTP in drinking water) factorial design for 12 weeks. METHODS: Masson's trichrome staining evaluated myocardial fibrosis in coronary vessels and surrounding myocardium. Whole blood specimens were counted for differentials. Spleen tumor necrosis factor-α (TNF-α) mRNA expression was determined by real-time RT-PCR. Data were analyzed by two-way analysis of variance (ANOVA) followed by mean separation procedures. RESULTS: After 12 weeks, lipopolysaccharide administration induced myocardial fibrosis in vessels and surrounding myocardium, spleen TNF-α mRNA expression, and leukocytes, while GTP supplementation in drinking water significantly averted such observation. CONCLUSIONS: GTP attenuates myocardial fibrosis through a suppression of chronic inflammation and innate immune responses.

Protective actions of green tea polyphenols and alfacalcidol on bone microstructure in female rats with chronic inflammation.

This study investigated the effects of green tea polyphenols (GTP) and alfacalcidol on bone microstructure and strength along with possible mechanisms in rats with chronic inflammation. A 12-week study using a 2 (no GTP vs. 0.5%, w/v GTP in drinking water)×2 (no alfacalcidol vs. 0.05 µg/kg alfacalcidol orally, 5×/week) factorial design was employed in lipopolysaccharide (LPS)-administered female rats. A group receiving placebo administration was used to compare with a group receiving LPS administration only to evaluate the effect of LPS. Changes in tibial and femoral microarchitecture and strength of femur were evaluated. Difference in expression of tumor necrosis factor-α (TNF-α) in proximal tibia using immunohistochemistry was examined. Compared to the placebo group, the LPS-administered-only group had significantly lower femoral mass, trabecular volume, thickness and number in proximal tibia and femur, and lower periosteal bone formation rate in tibial shafts but had significantly higher trabecular separation and osteoclast number in proximal tibia and eroded surface in endocortical tibial shafts. Both GTP and alfacalcidol reversed these LPS-induced detrimental changes in femur, proximal tibia and endocortical tibial shaft. Both GTP and alfacalcidol also significantly improved femoral strength, while significantly suppressed TNF-α expression in proximal tibia. There were significant interactions in femoral mass and strength, trabecular separation, osteoclast number and TNF-α expression in proximal tibia. A combination of both showed to sustain bone microarchitecture and strength. We conclude that a protective impact of GTP and alfacalcidol in bone microarchitecture during chronic inflammation may be due to a suppression of TNF-α.

Cytotoxic T cell immunity against the non-immunogenic, murine, hepatocellular carcinoma Hepa1-6 is directed towards the novel alternative form of macrophage colony stimulating factor.

Mouse Hepa1-6 hepatocellular carcinoma (HCC) cells were transduced with the membrane form of macrophage colony stimulating factor (mM-CSF). When mM-CSF transduced Hepa1-6 cells were injected subcutaneously into mice, these cells did not form tumors. The spleens of these immunized mice contained cytotoxic CD8+ T lymphocytes (CTL) that killed the unmodified Hepa1-6 cells. We show that the alternative form of macrophage colony stimulating factor (altM-CSF) induced CTL-mediated immunity against Hepa1-6 cells. AltM-CSF is restricted to the H-2D(b) allele. CTLs killed RMA-S cells loaded with exogenous altM-CSF peptide. Vaccination of mice with dendritic cells pulsed with the altM-CSF peptide stimulated anti-Hepa1-6 CTLs. Hyper-immunization of mice with mM-CSF Hepa1-6 cells showed inflammation of the liver and kidneys. Although altM-CSF was expressed within liver and kidney cells, its intensity was lower than Hepa1-6 cells. AltM-CSF was detected within the human HepG2 cell line. These studies suggest that altM-CSF may be a tumor antigen for HCC.

Comprehensive mutational analysis and mRNA isoform quantification of TP63 in normal and neoplastic human prostate cells.

BACKGROUND: The role of TP63 in cancer remains controversial since both oncogenic and tumor suppressive actions have been reported. p63 protein is found in the nuclei of basal cells of the normal prostate, yet it is absent in the vast majority of prostate cancer nuclei. Since a complex array of TP63 mRNA transcripts encode polypeptides with distinct functional properties, it is important to determine which forms are expressed in normal and prostate cancer tissue. METHODS: We used real-time RT-PCR to distinguish TP63 mRNA isoforms in prostate cancer cell lines (n = 7), and samples from prostate cancer patients. We sequenced all TP63 exons from 20 primary tumors, 20 metastases, 28 tumor xenografts, and 7 prostate cancer cell lines. RESULTS: TP63 mRNA isoforms were present in all tumors, albeit at levels lower than in normal prostate. The most abundant N-terminal variant was DeltaN; the most abundant C-terminal variant was the alpha form. The prostate tumor cell line CWR22Rv1 contained a single G to T substitution in exon 8 that is identical to a dominant-negative DNA binding inactivation mutation occurring in patients with a congenital TP63 deficiency syndrome. One patient tumor contained a somatic mutation in exon 11. CONCLUSIONS: The pattern of TP63 mRNA expression in normal prostate tissue is retained in reduced amounts in prostate cancer, and a potentially functional TP63 mutation was identified in one prostate tumor. Thus, if TP63 is a prostate cancer gene it likely functions as a tumor suppressor. Further study of the role of TP63 isoforms in regulating stem cell functions of normal and neoplastic prostate epithelial cells is needed. Prostate 69:559-569, 2009. (c) 2009 Wiley-Liss, Inc.