牛田 かおり

Adipocyte-cancer cell interactions promote tumor development and progression. Previously, we identified adipsin (CFD) and its downstream effector, hepatocyte growth factor (HGF), as adipokines that enhance adipocyte-breast cancer stem cell interactions. Here, we show that adipsin-dependent adipocyte maturation and the subsequent upregulation of HGF promote tumor invasion in breast cancers. Mature adipocytes, but not their precursors, significantly induced breast tumor cell migration and invasion in an adipsin expression-dependent manner. Promoters of tumor invasion, galectin 7 and matrix metalloproteinases, were significantly upregulated in cancer cells cocultured with mature adipocytes; meanwhile, their expression levels in cancer cells cocultured with adipocytes were reduced by adipsin knockout (Cfd KO) or a competitive inhibitor of CFD. Tumor growth and distant metastasis of mammary cancer cells were significantly suppressed when syngeneic mammary cancer cells were transplanted into Cfd KO mice. Histological analyses revealed reductions in capsular formation and tumor invasion at the cancer-adipocyte interface in the mammary tumors formed in Cfd KO mice. These findings indicate that adipsin-dependent adipocyte maturation may play an important role in adipocyte-cancer cell interaction and breast cancer progression.

<jats:title>Abstract</jats:title><jats:p>Epithelial–mesenchymal transition (EMT) plays a pivotal role in cancer metastasis and treatment resistance, which worsens prognosis. In phase III trials, eribulin improved overall survival in metastatic breast cancer (MBC) patients. In preclinical studies, eribulin suppressed EMT. However, clinical data on the use of eribulin for MBC patients are limited. In this exploratory, prospective study, we examined the effect of eribulin on EMT in MBC patients. Twenty‐two patients aged 44–82 years with recurrent breast cancer or MBC were treated with eribulin. Breast cancer tissue samples were obtained before treatment and on Day 15 ± 5 of the first cycle of eribulin treatment. EMT markers (E‐cadherin, claudin‐3, vimentin, and N‐cadherin) were analyzed using western blotting. EMT changes were evaluated based on the ratio of epithelial to mesenchymal markers before and after treatment in individual tumors. E‐cadherin/vimentin, claudin‐3/vimentin, E‐cadherin/N‐cadherin, and claudin‐3/N‐cadherin ratios were significantly higher after treatment (<jats:italic>p</jats:italic> = .007, <jats:italic>p</jats:italic> = .005, <jats:italic>p</jats:italic> = .006, and <jats:italic>p</jats:italic> = .011, respectively). Based on E‐cadherin/vimentin, 65.0% of tumors shifted to an epithelial phenotype, as compared to 66.7% based on claudin‐3/vimentin, 84.6% based on E‐cadherin/N‐cadherin, and 71.4% based on claudin‐3/N‐cadherin ratios. Thus, our results showed that eribulin suppressed EMT in breast cancer tissues.</jats:p>

<jats:title>Abstract</jats:title><jats:p>For pituitary regenerative medicine, the creation of a hypophyseal model in monkeys is necessary to conduct future preclinical studies; however, previous studies reported that hypophysectomy in monkeys is not always safe or satisfactory. This study aimed to create a hypophyseal dysfunction model in a cynomolgus monkey using a safer surgical technique and establish the protocol of pituitary hormone replacement therapy for this model. Surgical resection of the pituitary gland of a 7.8-year-old healthy adult cynomolgus male monkey weighing 5.45 kg was performed to create a hypophyseal dysfunction model for future regenerative studies. Endoscopic transoral transsphenoidal surgery was used to perform hypophysectomy under navigation support. These procedures were useful for confirming total removal of the pituitary gland without additional bone removal and preventing complications such as cerebrospinal fluid leakage. Total removal was confirmed by pathological examination and computed tomography. Hypopituitarism was verified with endocrinological examinations including stimulation tests. Postoperatively, the monkey’s general condition of hypopituitarism was treated with hormone replacement therapy, resulting in long-term survival. The success of a minimally invasive and safe surgical method and long-term survival indicate the creation of a hypophyseal dysfunction model in a cynomolgus monkey; hence, this protocol can be employed in the future.</jats:p>

<jats:title>Abstract</jats:title> <jats:sec> <jats:title /> <jats:p>Cancer-associated fibroblasts (CAF) constitute a major component of the tumor microenvironment. Recent observations in genetically engineered mouse models and clinical studies have suggested that there may exist at least two functionally different populations of CAFs, that is, cancer-promoting CAFs (pCAF) and cancer-restraining CAFs (rCAF). Although various pCAF markers have been identified, the identity of rCAFs remains unknown because of the lack of rCAF-specific marker(s). In this study, we found that Meflin, a glycosylphosphatidylinositol-anchored protein that is a marker of mesenchymal stromal/stem cells and maintains their undifferentiated state, is expressed by pancreatic stellate cells that are a source of CAFs in pancreatic ductal adenocarcinoma (PDAC). In situ hybridization analysis of 71 human PDAC tissues revealed that the infiltration of Meflin-positive CAFs correlated with favorable patient outcome. Consistent herewith, Meflin deficiency led to significant tumor progression with poorly differentiated histology in a PDAC mouse model. Similarly, genetic ablation of Meflin-positive CAFs resulted in poor differentiation of tumors in a syngeneic transplantation model. Conversely, delivery of a Meflin-expressing lentivirus into the tumor stroma or overexpression of Meflin in CAFs suppressed the growth of xenograft tumors. Lineage tracing revealed that Meflin-positive cells gave rise to α-smooth muscle actin-positive CAFs that are positive or negative for Meflin, suggesting a mechanism for generating CAF heterogeneity. Meflin deficiency or low expression resulted in straightened stromal collagen fibers, which represent a signature for aggressive tumors, in mouse or human PDAC tissues, respectively. Together, the data suggest that Meflin is a marker of rCAFs that suppress PDAC progression.</jats:p> </jats:sec> <jats:sec> <jats:title>Significance:</jats:title> <jats:p>Meflin marks and functionally contributes to a subset of cancer-associated fibroblasts that exert antitumoral effects.</jats:p> </jats:sec>

Malaria is caused by Plasmodium parasites and is one of the most life-threatening infectious diseases in humans. Infection can result in severe complications such as cerebral malaria, acute lung injury/acute respiratory distress syndrome, and acute renal injury. These complications are mainly caused by P. falciparum infection and are major causes of death associated with malaria. There are a few species of rodent-infective malaria parasites, and mice infected with such parasites are now widely used for screening candidate drugs and vaccines and for studying host immune responses and pathogenesis associated with disease-related complications. We found that mice of the NC/Jic strain infected with rodent malarial parasites exhibit distinctive disease-related complications such as cerebral malaria and nephrotic syndrome, in addition to a rapid increase in parasitemia. Here, we focus on the analysis of host genetic factors that affect malarial pathogenesis and describe the characteristic features, utility, and future prospects for exploitation of the NC/Jic strain as a novel mouse model for malaria research.

<jats:sec><jats:label /><jats:p>Embedding of tissue samples that maintains a desired orientation is critical for preparing sections suitable for diagnosis and study objectives. Methods to prepare tissue sections include: (i) paraffin embedding or snap‐freezing followed by microtome or cryostat sectioning; and (ii) agarose embedding followed by cutting on a vibrating microslicer. Although these methods are useful for routine laboratory work, preparation of small and fragile tissues such as mouse organs, small human biopsy samples, and cultured floating spheres is difficult and requires special skills. In particular, tissue specimen orientation can be lost during embedding in molds and subsequent sectioning. Here, we developed a method using low melting temperature (LM) gelatin either alone or mixed with agarose to preliminarily embed collected tissues that are either prefixed or unfixed, followed by conventional fixation, paraffin embedding, freezing, and sectioning. The advantage of the method is that the LM gelatin and its mixture with agarose can be handled at room temperature but quickly hardens at 4°C, which allows embedding, trimming, and arranging of small and fragile tissues in a desired orientation and are compatible with traditional stainings. Thus, this method can have various laboratory applications and can be modified according to the needs of each laboratory.</jats:p></jats:sec>

<jats:p> Summary </jats:p><jats:p> Tuft cells (TCs) are minor components of gastrointestinal epithelia, characterized by apical tufts and spool-shaped somas. The lack of reliable TC-markers has hindered the elucidation of its role. We developed site-specific and phosphorylation-status–specific antibodies against Girdin at tyrosine-1798 (pY1798) and found pY1798 immunostaining of mouse jejunum clearly depicted epithelial cells closely resembling TCs. This study aimed to validate pY1798 as a TC-marker. Double-fluorescence staining of intestines was performed with pY1798 and known TC-markers, for example, hematopoietic-prostaglandin-D-synthase (HPGDS), or doublecortin-like kinase 1 (DCLK1). Odds ratios (ORs) were calculated from cell counts to determine whether two markers were attracting (OR<1) or repelling (OR>1). In consequence, pY1798 signals strongly attracted those of known TC-markers. ORs for HPGDS in mouse stomach, small intestine, and colon were 0 for all, and 0.08 for DCLK1 in human small intestine. pY1798-positive cells in jejunum were distinct from other minor epithelial cells, including goblet, Paneth, and neuroendocrine cells. Thus, pY1798 was validated as a TC-marker. Interestingly, apoptosis inducers significantly increased relative TC frequencies despite the absence of proliferation at baseline. In conclusion, pY1798 is a novel TC-marker. Selective tyrosine phosphorylation and possible resistance to apoptosis inducers implied the activation of certain kinase(s) in TCs, which may become a clue to elucidate the enigmatic roles of TCs.  </jats:p>

<jats:p>In gastric cancer, the non‐canonical Wnt signaling pathway is activated by Wnt5a, which has a critical role in disease outcome. Previous studies have shown that Wnt5a mediates the expression of the extracellular matrix protein laminin γ2 through Rac and <jats:styled-content style="fixed-case">JNK</jats:styled-content> activation to promote gastric cancer progression. However, the mechanism of this regulatory pathway has not been completely addressed. The scaffold protein Dvl is a major component of the Wnt signaling pathway. Here, we show that Dvl‐associating protein with a high frequency of leucine residues (Daple) mediates Wnt5a‐induced laminin γ2 expression. Immunohistochemical analysis showed marked expression of Daple in advanced clinical stages of gastric cancer, where it highly correlated with Wnt5a/b and laminin γ2 expression, the depth of wall invasion, and the frequency of lymph node metastasis. In cultured cancer cells, Daple depletion led to the suppression of Wnt5a‐induced Rac and <jats:styled-content style="fixed-case">JNK</jats:styled-content> activation, laminin γ2 expression, and cell migration and invasion. Accordingly, Daple depletion also suppressed liver metastasis in a mouse xenograft model of gastric cancer. These results suggest that the non‐canonical Wnt signaling pathway contributes to gastric cancer progression at least in part via Daple, which provides a new therapeutic opportunity for the treatment of the disease.</jats:p>

<jats:title>Abstract</jats:title> <jats:p>PI3K–Akt signaling is critical for the development, progression, and metastasis of malignant tumors, but its role in the tumor microenvironment has been relatively little studied. Here, we report that the Akt substrate Girdin, an actin-binding protein that regulates cell migration, is expressed and activated by Akt phosphorylation in cancer-associated fibroblasts (CAF) and blood vessels within the tumor microenvironment. Lewis lung tumors grafted into mice defective in Akt-mediated Girdin phosphorylation (SA transgenic mice) exhibited a decrease in both CAF infiltration and tumor growth, compared with wild-type (WT) host control animals. Contrasting with the findings of other studies, we found that Akt-dependent phosphorylation of Girdin was not a rate-limiting step in the growth of endothelial cells. In addition, Lewis lung tumors displayed limited outgrowth when cotransplanted with CAF derived from tumor-bearing SA transgenic mice, compared with CAF derived from tumor-bearing WT mice. Collectively, our results revealed a role for Akt-mediated Girdin phosphorylation in CAF during tumor progression, highlighting the need to inhibit Akt function in both tumor cells and cells that comprise the tumor microenvironment. Cancer Res; 75(5); 813–23. ©2015 AACR.</jats:p>