M2 Phenotype Macrophages Colocalize with Schwann Cells in Human Dental Pulp.

Macrophages are immune cells with high plasticity that perform many functions related to tissue injury and repair. They are generally categorized as 2 functional phenotypes: M1 (proinflammatory) and M2 (anti-inflammatory and prohealing). To investigate the role of macrophages in human dental pulp, we examined the localization and distributional alterations of macrophages in healthy dental pulp as well as during the reparative process of pulp capping with mineral trioxide aggregate (MTA) and in cariously inflamed pulp of adult human teeth. We also quantified the populations of M1/M2 macrophages in healthy dental pulp by flow cytometric analysis. CD68+CD86+ cells (M1 phenotype) and CD68+CD163+ cells (M2 phenotype) were 2.11% ± 0.50% and 44.99% ± 2.22%, respectively, of 2.96% ± 0.41% CD68+ cells (pan-macrophages) in whole healthy dental pulp. Interestingly, M2 phenotype macrophages were associated with Schwann cells in healthy pulp, during mineralized bridge formation, and in pulp with carious infections in vivo. Furthermore, the M2 macrophages associated with Schwann cells expressed brain-derived neurotrophic factor (BDNF) under all in vivo conditions. Moreover, we found that plasma cells expressed BDNF. Coculture of Schwann cells isolated from human dental pulp and human monocytic cell line THP-1 showed that Schwann cells induced M2 phenotypic polarization of THP-1 cell-derived macrophages. The THP-1 macrophages that maintained contact with Schwann cells were stimulated, leading to elongation of their cell shape and expression of M2 phenotype marker CD163 in cocultures. In summary, we revealed the spatiotemporal localization of macrophages and potent induction of the M2 phenotype by Schwann cells in human dental pulp. M2 macrophages protect neural elements, whereas M1 cells promote neuronal destruction. Therefore, suppressing the neurodestructive M1 phenotype and maintaining the neuroprotective M2 phenotype of macrophages by Schwann cells may be critical for development of effective treatment strategies to maintain the viability of highly innervated dental pulp.

Changes in penile length after radical prostatectomy: effect of neoadjuvant androgen deprivation therapy.

Although reports have shown evidence for penile length (PL) shortening after radical prostatectomy (RP), the association between neoadjuvant androgen deprivation therapy (NADT) and PL after RP has yet to be determined. This study evaluates chronological changes in PL after NADT and RP. Stretched PLs (SPLs) of 143 patients, 41 of whom had undergone NADT, were measured before, 10 days after, and 1, 3, 6, 9, 12, 18, and 24 months after RP. Chronological erectile function and testosterone levels were then evaluated. SPL was shortest 10 days after RP in both the NADT (-) and NADT (+) groups and gradually recovered in length thereafter. SPL in the NADT (-) group was significantly longer than that in the NADT (+) group before RP. However, no significant differences in SPLs were found between both groups 6 months after RP. Although all subjects in the NADT (+) group had testosterone levels of <50 ng/dL before RP, such levels increased after RP. Before RP, the NADT (-) group was found to have significantly better erectile function than the NADT (+) group. However, differences in erectile function between the NADT (-) and NADT (+) groups after RP were not significant. This report is the first to show that among patients with prostate cancer, those who underwent NADT had greater PL recovery after RP than those who did not. Data regarding PL recovery after NADT and RP obtained in this study could be useful for patients with prostate cancer who plan to undergo such procedures.

Using surveillance data to simulate the impact of a hypothetical pre-entry tuberculosis screening programme in Japan.

BACKGROUND: Despite a growing burden of foreign-born tuberculosis (TB) patients, Japan does not currently practise pre-entry tuberculosis (TB) screening among foreign-born entrants. OBJECTIVE: To evaluate the impact of a hypothetical pre-entry TB screening programme among new foreign-born entrants into Japan. METHOD: Using publicly available sources, we estimated 1) the number of prevalent TB cases, defined as bacteriologically or clinically confirmed cases among new foreign-born entrants into Japan in 2015, and 2) the yield from a hypothetical pre-entry TB screening programme under three scenarios: Scenario A, in which screening would be required of all applicants intending to stay for 3 months; Scenario B, screening among applicants for visas for settlement purposes; and Scenario C, screening among student and technical intern visa applicants. RESULTS: The numbers of prevalent TB cases under Scenarios A, B and C were respectively 492, 54 and 248 out of a total of 328 791, 21 554 and 182 879 applicants, respectively 276, 29 and 137 of whom would be detected via the pre-entry screening programme, giving an yield of respectively 83.9, 134.5 and 74.9 per 100 000 screened under each scenario. CONCLUSION: The yield was the highest under Scenario B; however, the impact was greatest under Scenario A, in that it detected the greatest number of patients and thus contributed the most in reducing the burden of foreign-born TB cases in Japan.

Two-stage laparoscopic treatment for strangulated inguinal, femoral and obturator hernias: totally extraperitoneal repair followed by intestinal resection assisted by intraperitoneal laparoscopic exploration.

PURPOSE: Total extraperitoneal preperitoneal (TEP) repair is widely used for inguinal, femoral, or obturator hernia treatment. However, mesh repair is not often used for strangulated hernia treatment if intestinal resection is required because of the risk of postoperative mesh infection. Complete mesh repair is required for hernia treatment to prevent postoperative recurrence, particularly in patients with femoral or obturator hernia. CASES: We treated four patients with inguinocrural and obturator hernias (a 72-year-old male with a right indirect inguinal hernia; an 83-year-old female with a right obturator hernia; and 86- and 82-year-old females with femoral hernias) via a two-stage laparoscopic surgery. All patients were diagnosed with intestinal obstruction due to strangulated hernia. First, the incarcerated small intestine was released and then laparoscopically resected. Further, 8-24 days after the first surgery, bilateral TEP repairs were performed in all patients; the postoperative course was uneventful in all patients, and they were discharged 5-10 days after TEP repair. At present, no hernia recurrence has been reported in any patient. CONCLUSION: The two-stage laparoscopic treatment is safe for treatment of strangulated inguinal, femoral, and obturator hernias, and complete mesh repair via the TEP method can be performed in elderly patients to minimize the occurrence of mesh infection.

Cyclic mechanical pressure-loading alters epithelial homeostasis in a three-dimensional in vitro oral mucosa model: clinical implications for denture-wearers.

Denture-wearing affects the quality and quantity of epithelial cells in the underlying healthy oral mucosa. The physiologic mechanisms, however, are poorly understood. This study aimed to compare histologic changes and cellular responses of an epithelial cell layer to cyclic mechanical pressure-loading mimicking denture-wearing using an organotypic culture system to develop a three-dimensional in vitro oral mucosa model (3DOMM). Primary human oral keratinocytes and fibroblasts were serially grown in a monolayer culture, and cell viability was measured under continuous cyclic mechanical pressure (50 kPa) for 7 days (cycles of 60 min on, 20 s off to degas and inject air). Upon initiation of an air-liquid interface culture for epithelial stratification, the cyclic pressure, set to the mode above mentioned, was applied to the 3DOMMs for 7 days. Paraffin-embedded 3DOMMs were examined histologically and immunohistochemically. In the monolayer culture, the pressure did not affect the viability of oral keratinocytes or fibroblasts. Few histologic changes were observed in the epithelial layer of the control and pressure-loaded 3DOMMs. Immunohistochemical examination, however, revealed a significant decrease in Ki-67 labelling and an increase in filaggrin and involucrin expression in the suprabasal layer of the pressure-loaded 3DOMMs. Pressure-loading attenuated integrin ß1 expression and increased matrix metalloproteinase-9 activity. Incomplete deposition of laminin and type IV collagen beneath the basal cells was observed only in the pressure-loaded 3DOMM. Cyclic pressure-loading appeared to disrupt multiple functions of the basal cells in the 3DOMM, resulting in a predisposition towards terminal differentiation. Thus, denture-wearing could compromise oral epithelial homeostasis.

Zoledronic acid impairs re-epithelialization through down-regulation of integrin αvß6 and transforming growth factor beta signalling in a three-dimensional in vitro wound healing model.

This study examined the negative effects of zoledronic acid on the re-epithelialization of oral mucosa in a three-dimensional in vitro oral mucosa wound healing model. A living oral mucosa equivalent was constructed by seeding a mixture of primary human oral keratinocytes and fibroblasts, at a cell density of 1.5 × 10(5)cm(2) each, onto human cadaver dermis. This was cultured in a submerged condition in 1.2mM Ca(2+) EpiLife for 5 days, and then in an air-liquid interface for 14 days. The equivalent was wounded by excising a linear 2-mm-wide epithelial layer on day 8 and subsequently incubated with 10 µM zoledronic acid for an additional 11 days. Histological and immunohistochemical observations revealed zoledronic acid to significantly suppress the epithelial thickness and Ki-67-labelling index. Zoledronic acid also abolished integrin αvß6 expression, implying impaired keratinocyte migration. Zoledronic acid did not attenuate the total transforming growth factor beta 1 (TGF-ß1) production into the supernatant, but down-regulated TGF-ß receptor types I and II expression and Smad3 phosphorylation, as was also confirmed by immunofluorescence microscopy. This study therefore showed zoledronic acid to abrogate integrin αvß6 expression, cause the down-regulation of TGF-ß/Smad signalling in oral keratinocytes, and impair re-epithelialization, suggesting compromised oral mucosa homeostasis in patients receiving zoledronic acid.

Anti-androgen receptor ASC-J9 versus anti-androgens MDV3100 (Enzalutamide) or Casodex (Bicalutamide) leads to opposite effects on prostate cancer metastasis via differential modulation of macrophage infiltration and STAT3-CCL2 signaling.

Despite androgen deprivation therapy (ADT) suppression of prostate cancer (PCa) growth, its overall effects on PCa metastasis remain unclear. Using human (C4-2B/THP1) and mouse (TRAMP-C1/RAW264.7) PCa cells-macrophages co-culture systems, we found currently used anti-androgens, MDV3100 (enzalutamide) or Casodex (bicalutamide), promoted macrophage migration to PCa cells that consequently led to enhanced PCa cell invasion. In contrast, the AR degradation enhancer, ASC-J9, suppressed both macrophage migration and subsequent PCa cell invasion. Mechanism dissection showed that Casodex/MDV3100 reduced the AR-mediated PIAS3 expression and enhanced the pSTAT3-CCL2 pathway. Addition of CCR2 antagonist reversed the Casodex/MDV3100-induced macrophage migration and PCa cell invasion. In contrast, ASC-J9 could regulate pSTAT3-CCL2 signaling using two pathways: an AR-dependent pathway via inhibiting PIAS3 expression and an AR-independent pathway via direct inhibition of the STAT3 phosphorylation/activation. These findings were confirmed in the in vivo mouse model with orthotopically injected TRAMP-C1 cells. Together, these results may raise the potential concern about the currently used ADT with anti-androgens that promotes PCa metastasis and may provide some new and better therapeutic strategies using ASC-J9 alone or a combinational therapy that simultaneously targets androgens/AR signaling and PIAS3-pSTAT3-CCL2 signaling to better battle PCa growth and metastasis at castration-resistant stage.

Nuclear hormone receptor signals as new therapeutic targets for urothelial carcinoma.

Unlike prostate and breast cancers, urothelial carcinoma of the urinary bladder is not yet considered as an endocrine-related neoplasm, and hormonal therapy for bladder cancer remains experimental. Nonetheless, there is increasing evidence indicating that nuclear hormone receptor signals are implicated in the development and progression of bladder cancer. Androgen-mediated androgen receptor (AR) signals have been convincingly shown to induce bladder tumorigenesis. Androgens also promote the growth of AR-positive bladder cancer cells, although it is controversial whether AR plays a dominant role in bladder cancer progression. Both stimulatory and inhibitory functions of estrogen receptor signals in bladder cancer have been reported. Various studies have also demonstrated the involvement of other nuclear receptors, including progesterone receptor, glucocorticoid receptor, vitamin D receptor, and retinoid receptors, as well as some orphan receptors, in bladder cancer. This review summarizes and discusses available data suggesting the modulation of bladder carcinogenesis and cancer progression via nuclear hormone receptor signaling pathways. These pathways have the potential to be an extremely important area of bladder cancer research, leading to the development of effective chemopreventive/therapeutic approaches, using hormonal manipulation. Considerable uncertainty remains regarding the selection of patients who are likely to benefit from hormonal therapy and optimal options for the treatment.

Risedronate prevents persistent bone loss in prostate cancer patients treated with androgen deprivation therapy: results of a 2-year follow-up study.

Androgen deprivation therapy (ADT) for prostate cancer (PCa) causes bone loss. Although we reported previously that risedronate significantly recovers bone mineral density (BMD) for up to 12 months, there have been no reports with longer follow-up periods to date. This study extended our earlier series extending the follow-up period to 24 months. Eligible patients had histologically confirmed PCa without lumbar spine metastasis and underwent ADT. Lumbar spine BMD, urinary deoxypyridinoline (uDPD) and serum bone alkaline phosphatase were measured at 6, 12 and 24 months. Among the total of 96 patients, we analyzed 26 and 18 patients in risedronate administration and control groups, respectively. BMD relative to the young adult mean ratio, uDPD and serum bone alkaline phosphatase of the risedronate administration group recovered significantly after 24 months compared with the control group (P<0.0001, P=0.0001, and P<0.0001, respectively). Transient blurred vision, malaise and vertigo were observed in 1 patient each among the 46 patients treated with risedronate within 28 days after first administration. Oral administration of risedronate is safe and effective for the recovery of ADT-induced bone loss in PCa patients even at 24 months after commencement of treatment.

Blockade of Th1 chemokine receptors ameliorates pulmonary granulomatosis in mice.

Sarcoidosis is a granulomatous disease of unknown aetiology. We identified immunological targets for the treatment of pulmonary granulomatosis using a murine model generated with Propionibacterium acnes. Sensitisation and challenge using heat-killed P. acnes and dendritic cells (DCs) were performed to produce pulmonary granulomatosis in C57BL/6 mice. Immunological analyses using ELISA as well as cDNA microarray analysis were used to search for cytokines or chemokines associated with the formation of granulomas in the lungs. Co-administration of P. acnes and DCs reproducibly induced the formation of pulmonary granulomas, which resembled sarcoid granulomas. The cDNA microarray assay demonstrated that the gene expression of CXCL9 and CXCL10, ligands for CXCR3, and of CCL4, a ligand for CCR5, was strongly upregulated during granulomatosis. ELISA confirmed that levels of CXCL9 and CXCL10 as well as T-helper (Th)1 cytokines and chemokines including tumour necrosis factor-α and interferon-γ were elevated in bronchoalveolar lavage fluid (BALF). The blockade of Th1 chemokine receptors using TAK-779, a dual blocker for CXCR3 and CCR5, led to reduced numbers of CXCR3+CD4+ and CCR5+CD4+ T-cells in BALF. Furthermore, administration of TAK-779 ameliorated the granulomatosis. The targeted inhibition of Th1 chemokines might be useful for inhibiting Th1-biased granulomatous diseases, including sarcoidosis.

Enhancement of osteoclastogenic activity in osteolytic prostate cancer cells by physical contact with osteoblasts.

BACKGROUND: The interaction between prostate cancer cells and osteoblasts is critical for the development of bone metastasis. Metastatic cancer cells may physically contact osteoblasts in the bone microenvironment; however, the biological significance of this interaction is not fully understood. METHODS: Human prostate cancer cells (the osteolytic cell line PC-3 and the osteoblastic cell line MDA-PCa 2b) and human osteoblasts (hFOB1.19) were cocultured under two different conditions (bilayer and contact conditions). Differential gene expression profiles of prostate cancer cells were then investigated using microarray analysis. Differentially expressed genes were analysed using RT-PCR and western blotting, and the effect of anti-cadherin neutralising antibodies on their expression was assayed. The osteoclastogenic activity of cells grown under these different conditions was also investigated using an in vitro assay. RESULTS: When PC-3 or MDA-PCa 2b cells were cocultured with hFOB1.19 cells under contact conditions, the expression of eight genes was upregulated and that of one gene was downregulated in PC-3 cells compared with gene expression in bilayer culture. No differentially expressed genes were detected in MDA-PCa 2b cells. Four of the eight upregulated genes (interleukin-1ß (IL-1ß), cyclooxygenase-2 (COX-2), IL-6 and the third component of complement (C3)) have already been reported to participate in osteoclastogenesis. Indeed, a cell lysate of PC-3 cells grown under contact coculture conditions significantly enhanced osteoclastogenesis in vitro (P<0.005). neutralisation of cadherin-11 with a specific antibody inhibited upregulation of COX-2 and C3 mRNA in PC-3 cells. In contrast, neutralisation of N-cadherin induced upregulation of COX-2 mRNA. CONCLUSION: Physical contact between osteolytic prostate cancer cells and osteoblasts may upregulate osteoclastogenesis-related gene expression in prostate cancer cells and enhance osteoclastogenesis. Additionally, cadherin-11 and N-cadherin are involved in this process. These data provide evidence supporting new therapies of prostate cancer bone metastasis that target direct cancer-cell-osteoblast cell-cell contact.

Depressive symptoms, not completing a depression screening questionnaire, and risk of poor compliance with regular primary care visits in patients with type 2 diabetes: the Japan Diabetes Outcome Intervention Trial 2 (J-DOIT2) study group.

OBJECTIVES: To explore the association between depressive symptoms as measured by the Center for Epidemiologic Studies Depression Scale or not completing the questionnaire and subsequent risk of poor compliance with regular visits to primary care physician in patients with type 2 diabetes. METHODS: Using data from patients with type 2 diabetes who participated in the Japan Diabetes Outcome Intervention Trial 2 (J-DOIT2) Pilot Study, which was conducted at primary care settings, we examined the association between depressive symptoms or not completing the questionnaire and risk of poor compliance with regular visits as an event. RESULTS: Among 1 584 patients who participated in the J-DOIT2 Pilot Study, we excluded 140 who did not meet inclusion criteria or who declined participation after randomization, leaving 1 444 for entry in the present analysis. During 1 409 person-years of follow-up (median 1 year), 90 events were observed (incidence rate 63.9/1 000 person-years). The multivariable-adjusted hazard ratio of poor compliance with regular visits in those having depressive symptoms was 1.23 (95% CI: 0.46-3.31). In contrast, the multivariable-adjusted hazard ratio of poor compliance in those not completing the questionnaire was 2.26 (95% CI: 1.94-2.63). CONCLUSION: Not completing a questionnaire was significantly associated with an increased risk of poor compliance with the maintenance of regular visits to a primary care physician in patients with type 2 diabetes. Patients who do not comply with questionnaire surveys require increased attention to ensure their compliance with regular visits, and thereby ensure better diabetes outcomes.

Formaldehyde measurements in residential indoor air using a developed sensor element in the Kanto area of Japan.

UNLABELLED: We undertook this to determine the formaldehyde concentration in Japanese houses and the relationship between formaldehyde levels and the age and temperature of a house using a sensor element that we developed for time-integrated measurements of formaldehyde concentration in actual environments. We evaluated the correlation between the formaldehyde concentration estimated by the dinitrophenylhydrazine (DNPH)-derivatization method and that obtained with our sensor element. We found a linear relationship between the two results indicating that reliable measurements can be performed using the developed sensor element in actual environments. The indoor concentration of formaldehyde was determined in a study of 34 homes in the Kanto area of Japan, between September 28 and October 27, 2007. We obtained the highest formaldehyde concentrations of 92 ± 15 µg/m(3) for apartments 0-2 years after their renovation, and a simple linear relationship was found between formaldehyde concentration and the age of the apartment. We also found that the formaldehyde concentration in a room containing furniture increased by 10% when the temperature increased by 1°C. PRACTICAL IMPLICATIONS: This study contributed to the measurements of indoor formaldehyde levels. We have used a newly developed sensor for time-integrated measurements of formaldehyde concentrations. This sensor does not need a power supply during exposure to air, and does not need special skills to use. This research showed that homeowners successfully deployed the sensor at the desired place and desired period in their house by themselves. Formaldehyde is emitted by various off-gassing sources, such as furniture. Therefore, for example, homeowners may want to measure the change of formaldehyde levels in their house before and after installing new furniture. This sensor may also be deployed by occupants to reduce the cost of a large-scale exposure assessment study.

Development of an in vitro model for radiation-induced effects on oral keratinocytes.

Changes in epithelial cell activity and the production of pro-inflammatory cytokines were examined utilizing an organotypic culture system as an in vitro model to study the effects of radiation on oral keratinocytes to simulate what is thought to occur in radiation-induced oral mucositis. Monolayer cultures of oral keratinocyte were irradiated by varying the dose. Cell injury was assessed using a colony forming efficiency (CFE) assay. Third passage oral keratinocytes were seeded onto AlloDerm to form a 3D construct of an ex vivo produced oral mucosa equivalent (EVPOME) which was irradiated with 0, 1, 3 and 8Gy. Formalin-fixed sections of the EVPOME were used for histology and immunohistochemistry to examine proliferative capacity. Epithelial cell viability of EVPOME was measured by MTT assay. Spent culture medium was used to determine post-radiation pro-inflammatory cytokine production. Basal cells became more swollen and pyknotic as radiation increased, implying loss of cell viability also determined by MTT assay. The number of Ki-67 immunopositive cells and CFE showed negative correlation with radiation, indicating loss of cell proliferative capacity. The production of pro-inflammatory cytokines, IL-1alpha and IL-8, tended to increase in a radiation dose dependent manner. The EVPOME lacking submucosal cellular components was a useful model.

Pharmacological retention of oral mucosa progenitor/stem cells.

Oral mucosa progenitor/stem cells reside as a small-sized cell population that eventually differentiates concurrently with an increase in cell size. Activation of the mammalian target of rapamycin (mTOR) leads to an increase in cell size. We hypothesized that rapamycin, a specific inhibitor of mTOR, will maintain primary human oral keratinocytes as a small-sized, undifferentiated cell population capable of retaining their proliferative capacity. Primary, rapamycin-treated (2 nM, 20 nM) oral keratinocytes showed a diminished cell size that correlated with a higher clonogenicity, a longer-term proliferative potential, and a slower cycling cell population concurrent with decreased expression of a differentiation marker when compared with untreated cells. Only the 2-nM rapamycin-treated oral keratinocytes maintained their ability to regenerate oral mucosa in vitro after 15 weeks of culture. Rapamycin, a Food and Drug Administration-approved drug, may have applicability for use in creating a highly proliferative cell population for use in regenerative medicine.

Usefulness of real-time tissue elastography for detecting lymph-node metastases in squamous cell carcinoma.

We report a case of invasive SCC arising from multiple lesions of Bowen's disease with right inguinal lymph-node metastasis. Assessment of superficial lymph-node involvement by real-time tissue elastography before surgery was found to be more useful than other noninvasive conventional methods. Histologically, the metastatic tumour cells were located asymmetrically in a small section of the cortical area of the right node, and this result was comparable with the elastographic findings. Additionally, we found that the presence of an asymmetrical cortical area with high elasticity should be included in the determination of metastatic involvement in small lymph nodes. It has high predictive values in the differentiation of benign and malignant superficial lymph nodes in patients with clinically node-negative skin cancer. More cases are needed to validate this efficiency in differentiating benign from malignant lymph-node status, but if confirmed, it may have an important role in the diagnosis of high-risk cutaneous squamous cell carcinoma.

The expression and production of vascular endothelial growth factor in oral mucosa equivalents.

Angiogenesis is anticipated during wound healing. Vascular endothelial growth factor (VEGF) is a potent direct angiogenic factor that stimulates endothelial cell migration and proliferation in vitro and angiogenesis in vivo. Ex vivo produced oral mucosa equivalent (EVPOME) grafts have been reported to promote re-vascularization in the underlying tissue after grafting. The aim of this study was to evaluating the following: VEGF mRNA and its protein expression in EVPOME grafts, the protein levels in conditioned media produced by EVPOMEs, and the ability of VEGF to stimulate the growth of microvascular endothelial cells. VEGF mRNA expression and immunoreaction were found in basal and suprabasal layers. VEGF secreted by EVPOME was detected throughout the period of manufacture of the grafts, and became elevated for the first week at an air-liquid interface. Both EVPOME-conditioned media and a medium containing 10ng/mL recombinant human VEGF induced endothelial cell proliferation, while neutralization of VEGF by an antibody blocked cell growth. These results suggest that VEGF secreted by EVPOME grafts might assist initial vascularization after grafting, which is critical to subsequent graft survival.