Fluctuation of CD9/SOX2-positive cell populations during the turnover of GH- and TSH-producing cells in the adult anterior pituitary gland.

The adenohypophysis is comprised of the anterior and intermediate lobes (AL and IL, respectively). Cluster of differentiation 9 (CD9)- and sex-determining region Y-box 2 (SOX2)-positive cells are stem/progenitor hormone-producing cells in the AL. They are located in the marginal cell layer (MCL) facing Rathke's cleft between the AL and IL (primary niche) and the parenchyma of the AL (secondary niche). We previously showed that, in rats, CD9/SOX2-positive cells in the IL side of the MCL (IL-side MCL) migrate to the AL side (AL-side MCL) and differentiate into prolactin-producing cells (PRL cells) in the AL parenchyma during pregnancy, lactation, and diethylstilbestrol treatment, all of which increase PRL cell turnover. This study examined the changes in CD9/SOX2-positive stem/progenitor cell niches and their proportions by manipulating the turnover of growth hormone (GH)- and thyroid-stimulating hormone (TSH)-producing cells (GH and TSH cells, respectively), which are Pit1 lineage cells, as well as PRL cells. After induction, the isolated CD9/SOX2-positive cells from the IL-side MCL formed spheres and differentiated into GH and TSH cells. We also observed an increased GH cell proportion upon treatment with GH-releasing hormone and recovery from continuous stress and an increased TSH cell proportion upon propylthiouracil treatment, concomitant with alterations in the proportion of CD9/SOX2-positive cells in the primary and secondary niches. These findings suggest that CD9/SOX2-positive cells have the potential to supply GH and TSH when an increase in GH and TSH cell populations is required in the adult pituitary gland.

Antiphase synchrony increases perceived entitativity and uniqueness: A joint hand-clapping task.

In- and antiphase are the dominant patterns identified in the study of synchrony in relative phases. Many previous studies have focused on in-phase synchrony and compared it to asynchrony, but antiphase synchrony has yet not been the subject of much research attention. The limited findings on antiphase synchrony suggest that its role or nature is unclear or unstable in human interaction. To account for this factor, this study examined the possibility that antiphase synchrony simultaneously induced perceived entitativity and uniqueness. The results of an experiment employing a joint hand-clapping task supported this prediction. Further, the elevated feeling of uniqueness in those who experienced antiphase synchrony may have increased the self-other overlap for those who felt oneness with their partner, but it decreased overlap for those who did not. The theoretical implications for synchrony literature are discussed.

Using Virtual Reality to Improve the Quality of Life of Older Adults with Cognitive Impairments and their Family Members who Live at a Distance.

This study examines whether using virtual reality (VR) with older adults with mild cognitive impairment (MCI) or mild to moderate dementia with a family member who lives at a distance can improve the quality of life of the older adult and the family member. Twenty-one older adults in a senior living community and a family member (who participated in the VR with the older adult from a distance) engaged in a baseline telephone call, followed by three weekly VR sessions. The VR was associated with improvements in older adults' affect and stress, relationship with their family member, and overall quality of life, compared to baseline. Family members' negative affect, depressive symptoms, and caregiver burden also decreased and their mental health improved after using the VR, compared to baseline. Using the VR, however, did not change their relationship with the older adult. In addition, older adults and family members who experienced the VR sessions as more socially engaging reported better psychological and relational well-being, with older adults also experiencing greater improvements in overall quality of life. Finally, preliminary results suggest that older adults with dementia and their family members might benefit even more from using the VR than older adults with MCI and their family members.

Nonverbal Synchrony in Technology-Mediated Interviews: A Cross-Cultural Study.

Technology-mediated communication has changed the way we interact. Since the onset of the COVID-19 pandemic in March 2020, this trend became even more pronounced. Media interviews are no exception. Yet, studies on nonverbal behaviors, especially nonverbal synchrony during such mediated settings, have been scarce. To fill the research gap, this study investigated synchronized patterns between interview hosts' and guests' facial emotional displays and upper body movement during mediated interviews recorded in the countries in Western (mainly the US, with the addition of the UK) and Eastern cultures (Japan). The interviews were categorized into information- or entertainment-driven interviews, depending on the social attributes of the guest. The time series of the valence in facial displays and upper body movement was automatedly measured using FaceReader and Motion Energy Analysis software, respectively, which was analyzed in terms of simultaneous movements, a primary component of synchrony. As predicted, facial synchrony was more prevalent in information-driven interviews, supporting the motivational and strategic account of synchrony. In addition, female-hosted interviews had a higher degree of synchrony, especially in information-driven interviews. Similar patterns were seen in movement synchrony, although not significant. This study is the first evidence of synchrony in technology-mediated interviews in which a host and a guest appear on split-screen to inform or entertain audiences. However, no cultural differences in synchrony were observed. Situational demands in front of the interactants and the goal-driven nature of communication seemed to play a more prominent role than cultural differences in nonverbal synchrony. Supplementary Information: The online version contains supplementary material available at 10.1007/s10919-022-00416-3.

Differentiation of stem progenitor CD9/SOX2-positive cells is promoted with increased prolactin-producing and endothelial cells in the pituitary.

Sex-determining region Y-box 2 (SOX2)-positive cells are stem/progenitor cells in the adenohypophysis, comprising the anterior and intermediate lobes (AL and IL, respectively). The cells are located in the marginal cell layer (MCL) facing Rathke's cleft (primary niche) and the parenchyma of the AL (secondary niche). We previously demonstrated in vitro that the tetraspanin superfamily CD9 and SOX2 double-positive (CD9/SOX2-positive) cells in the IL-side MCL migrate to the AL side and differentiate into hormone-producing and endothelial cells in the AL parenchyma. Here, we performed in vivo studies to evaluate the role of IL-side CD9/SOX2-positive cells in pregnancy, lactation, and treatment with diethylstilbestrol (DES; an estrogen analog) when an increased population of prolactin (PRL) cells was observed in the AL of the rat pituitary. The proportions of CD9/SOX2-, CD9/Ki67-, and PRL/TUNEL-positive cells decreased in the primary and secondary niches during pregnancy and DES treatment. In contrast, the number of CD9/PRL-positive cells increased in the AL-side MCL and AL parenchyma during pregnancy and during DES treatment. The proportion of PRL/Ki67-positive cells increased in the AL-side MCL and AL parenchyma in response to DES treatment. Next, we isolated CD9-positive cells from the IL-side MCL using an anti-CD9 antibody. During cell culture, the cells formed free-floating three-dimensional clusters (pituispheres). Furthermore, CD9-positive cells in the pituisphere differentiated into PRL cells, and their differentiation potential was promoted by DES. These findings suggest that CD9/SOX2-positive cells in the IL-side MCL may act as adult stem cells in the AL parenchyma that supply PRL cells under the influence of estrogen.

Expression and localization of tight junction-related proteins in adult rat pituitary stem/progenitor cell niches.

Pituitary endocrine cells are supplied by Sox2-expressing stem/progenitor cells in the anterior lobe of the adult pituitary gland. These SOX2-positive cells are maintained in two types of microenvironments (niches): the marginal cell layer (MCL)-niche and the parenchymal-niche. Recently, we isolated dense SOX2-positive cell clusters from the parenchymal-niche by taking advantage of their resistance to protease treatment as parenchymal stem/progenitor cell (PS)-clusters. In the present study, by analyzing these isolated PS-clusters, we attempted to identify novel structural characteristics of pituitary stem/progenitor cell niches. Quantitative real-time PCR showed that tight junction-related genes were distinctly expressed in the isolated PS-clusters. Immunocytostaining showed that the tight junction molecules, ZO-1 and occludin, were localized in the apical membrane facing the pseudo-follicle-like structure of the isolated PS-clusters regardless of the expression of S100ß, which distinguishes the sub-population of SOX2-positive cells. Furthermore, immunohistochemistry of the pituitary glands of adult rats clearly demonstrated that ZO-1 and occludin were densely present in the parenchymal-niche encircling the pseudo-follicle, while they were observed in the apical membrane in the MCL-niche facing the residual lumen. Collectively, these tight junction-related proteins might be involved in the architecture and maintenance of the plasticity of pituitary stem/progenitor cell niches.

The multiciliated cells in Rathke's cleft express CYP26A1 and respond to retinoic acid in the pituitary.

The adenohypophysis consists of the anterior and intermediate lobes (AL and IL). The marginal cell layer (MCL), including the ventral region of the IL and the dorsal region of the AL lining the Rathke's cleft, acts as the primary stem/progenitor cell niches in adult adenohypophysis. The cells of the MCL on the IL side consisted of cluster of differentiation 9 (CD9)-positive stem/progenitor cells with or without motile cilia. However, any additional cellular properties of multiciliated CD9-positive cells are not known. The present study aimed to identify the character of the multiciliated cells in stem cell niche of the pituitary gland. We observed the fine structure of the multiciliated cells in the MCL of male Wistar rats at an early stage after birth and in adulthood (P60) using scanning electron microscopy. Since the previous study showed that the MCL cells of adult rats synthesize retinoic acid (RA), the present study determined whether the multiciliated cells are involved in RA regulation by the expression of retinal aldehyde dehydrogenase 1 (RALDH1) and CYP26A1, an enzyme synthesizing and degrading RA, respectively. Results showed that 96% of multiciliated cells in adult male rats expressed CYP26A1, while 60% expressed RALDH1. Furthermore, the isolated CD9-positive cells from the IL side MCL responded to RA and activated the degradation system of RA by increasing Cyp26a1 expression. These findings indicated that multiciliated cells are involved in RA metabolism in the MCL. Our observations provide novel insights regarding the stem cell niche of the adult pituitary.

Empathic accuracy and interpersonal coordination: behavior matching can enhance accuracy but interactional synchrony may not.

The study considered two major facets of interpersonal coordination, namely, behavior matching (posture mirroring) and interactional synchrony, and investigated whether interpersonal coordination enhanced empathic accuracy. Interactional synchrony was further classified into simultaneous movement and interaction rhythms. Participants engaged in an eight-minute conversation with a same-gender unacquainted partner and an empathic accuracy task. Each participant viewed the interaction video and reported their thoughts and feelings at pre-determined points of time. Afterward, they rewatched the video and inferred the thoughts and feelings of their partners. The study employed OpenPose, 2D pose estimation software of human body, to quantify posture and bodily movement, which were used to determine coordination. The results indicated that behavior matching was positively associated with empathic accuracy, whereas rhythmic convergence in synchrony was negatively associated with accuracy in female dyads. The additional analysis explored the temporal relationship between coordination and accuracy, which indicated a cause-effect tendency during interactions.

Testing the Feasibility of Virtual Reality With Older Adults With Cognitive Impairments and Their Family Members Who Live at a Distance.

BACKGROUND AND OBJECTIVES: This study tests the feasibility of using virtual reality (VR) with older adults with mild cognitive impairment (MCI) or mild-to-moderate dementia with a family member who lives at a distance. RESEARCH DESIGN AND METHODS: 21 residents in a senior living community and a family member (who participated in the VR with the older adult from a distance) engaged in a baseline telephone call, followed by 3 weekly VR sessions. RESULTS: Residents and family members alike found the VR safe, extremely enjoyable, and easy to use. The VR was also acceptable and highly satisfying for residents with MCI and dementia. Human and automated coding revealed that residents were more conversationally and behaviorally engaged with their family member in the VR sessions compared to the baseline telephone call and in the VR sessions that used reminiscence therapy. The results also illustrate the importance of using multiple methods to assess engagement. Residents with dementia reported greater immersion in the VR than residents with MCI. However, the automated coding indicated that residents with MCI were more kinesically engaged while using the VR than residents with dementia. DISCUSSION AND IMPLICATIONS: Combining networking and livestreaming features in a single VR platform can allow older adults in senior living communities to still travel, relive their past, and engage fully with life with their family members, despite geographical separation and physical and cognitive challenges.

Insight into the Characteristics of Novel Desmin-Immunopositive Perivascular Cells of the Anterior Pituitary Gland Using Transmission and Focused Ion Beam Scanning Electron Microscopy.

Recently, another new cell type was found in the perivascular space called a novel desmin-immunopositive perivascular (DIP) cell. However, the differences between this novel cell type and other nonhormone-producing cells have not been clarified. Therefore, we introduced several microscopic techniques to gain insight into the morphological characteristics of this novel DIP cell. We succeeded in identifying novel DIP cells under light microscopy using desmin immunocryosection, combining resin embedding blocks and immunoelectron microscopy. In conventional transmission electron microscopy, folliculostellate cells, capsular fibroblasts, macrophages, and pericytes presented a flat cisternae of rough endoplasmic reticulum, whereas those of novel DIP cells had a dilated pattern. The number of novel DIP cells was greatest in the intact rats, though nearly disappeared under prolactinoma conditions. Additionally, focused ion beam scanning electron microscopy showed that these novel DIP cells had multidirectional processes and some processes reached the capillary, but these processes did not tightly wrap the vessel, as is the case with pericytes. Interestingly, we found that the rough endoplasmic reticulum was globular and dispersed throughout the cytoplasmic processes after three-dimensional reconstruction. This study clearly confirms that novel DIP cells are a new cell type in the rat anterior pituitary gland, with unique characteristics.

Liver-Directed AAV8 Booster Vaccine Expressing Plasmodium falciparum Antigen Following Adenovirus Vaccine Priming Elicits Sterile Protection in a Murine Model.

Hepatocyte infection by malaria sporozoites is a bottleneck in the life-cycle of Plasmodium spp. including P. falciparum, which causes the most lethal form of malaria. Therefore, developing an effective vaccine capable of inducing the strong humoral and cellular immune responses necessary to block the pre-erythrocytic stage has potential to overcome the spatiotemporal hindrances pertaining to parasite biology and hepatic microanatomy. We recently showed that when combined with a human adenovirus type 5 (AdHu5)-priming vaccine, adeno-associated virus serotype 1 (AAV1) is a potent booster malaria vaccine vector capable of inducing strong and long-lasting protective immune responses in a rodent malaria model. Here, we evaluated the protective efficacy of a hepatotropic virus, adeno-associated virus serotype 8 (AAV8), as a booster vector because it can deliver a transgene potently and rapidly to the liver, the organ malaria sporozoites initially infect and multiply in following sporozoite injection by the bite of an infected mosquito. We first generated an AAV8-vectored vaccine expressing P. falciparum circumsporozoite protein (PfCSP). Intravenous (i.v.) administration of AAV8-PfCSP to mice initially primed with AdHu5-PfCSP resulted in a hepatocyte transduction rate ~2.5 times above that seen with intramuscular (i.m.) administration. This immunization regimen provided a better protection rate (100% sterile protection) than that of the i.m. AdHu5-prime/i.m. AAV8-boost regimen (60%, p < 0.05), i.m. AdHu5-prime/i.v. AAV1-boost (78%), or i.m. AdHu5-prime/i.m. AAV1-boost (80%) against challenge with transgenic PfCSP-expressing P. berghei sporozoites. Compared with the i.m. AdHu5-prime/i.v. AAV1-boost regimen, three other regimens induced higher levels of PfCSP-specific humoral immune responses. Importantly, a single i.v. dose of AAV8-PfCSP recruited CD8+ T cells, especially resident memory CD8+ T cells, in the liver. These data suggest that boost with i.v. AAV8-PfCSP can improve humoral and cellular immune responses in BALB/c mice. Therefore, this regimen holds great promise as a next-generation platform for the development of an effective malaria vaccine.

CD9-positive cells in the intermediate lobe of the pituitary gland are important supplier for prolactin-producing cells in the anterior lobe.

A supply of hormone-producing cells from stem/progenitor cells is critical to sustain the endocrine activity of the pituitary gland. In the adenohypophysis composing the anterior and intermediate lobe (AL and IL, respectively), stem/progenitor cells expressing sex-determining region Y-box 2 (SOX2) and S100ß are located in the marginal cell layer (MCL) facing Rathke's cleft (primary niche) and the parenchyma of the AL (secondary niche). Our previous studies using mice and rats indicated that the tetraspanin superfamily CD9 and CD81 are expressed in S100ß/SOX2-positive cells of primary and secondary niches (named CD9/CD81/S100ß/SOX2-positive cell), and the cells located in the AL-side niches exhibit plasticity and multipotency. However, it is unclear whether CD9/CD81/S100ß/SOX2-positive cells in the IL-side primary niche are stem/progenitor cells for the AL or IL. Here, we successfully isolated pure CD9/CD81/S100ß/SOX2-positive cells from the IL-side primary niche. They had a higher level of S100ß and SOX2 mRNA and a greater pituisphere forming capacity than those of CD9/CD81/S100ß/SOX2-positive cells isolated from the AL. They also had capacity to differentiate into all types of adenohypophyseal hormone-producing cells, concomitantly with the loss of CD9 expression. Loss of CD9 and CD81 function in CD9/CD81/S100ß/SOX2-positive cells by siRNA treatment impaired prolactin cell differentiation. Consistently, in the pituitary gland of CD9/CD81 double knockout mice, dysgenesis of the MCL and a lower population of prolactin cells were observed. These results suggest that the CD9/CD81/S100ß/SOX2-positive cells in the MCL of the IL-side are potential suppliers of adult core stem cells in the AL.

Comparison of alemtuzumab, anti-thymocyte globulin, and post-transplant cyclophosphamide for graft-versus-host disease and graft-versus-leukemia in murine models.

Graft-versus-host disease is a major complication after allogeneic hematopoietic stem cell transplantation for hematological malignancies. Immunosuppressive drugs, such as anti-thymocyte globulin, alemtuzumab, and post-transplant cyclophosphamide, have been used to prevent graft-versus-host disease in HLA-mismatched haploidentical hematopoietic stem cell transplantation. Here, we investigated whether these drugs could ameliorate graft-versus-host disease without diminishing the graft-versus-leukemia effect by using a xenogeneic transplanted graft-versus-host disease/graft-versus-leukemia model. Anti-thymocyte globulin treatment diminished graft-versus-host disease symptoms, completely depleted the infiltration of inflammatory cells in the liver and intestine, and led to prolonged survival. By contrast, improvement after post-transplant cyclophosphamide treatment remained minimal. Alemtuzumab treatment modestly prolonged survival despite an apparent decrease of Tregs. In the graft-versus-leukemia model, 1.5 to 2.0 mg/kg of anti-thymocyte globulin and 0.6 to 0.9 mg/kg of alemtuzumab reduced graft-versus-host disease with minimal loss of graft-versus-leukemia effect. Mice treated with 400 mg/kg of post-transplant cyclophosphamide did not develop graft-versus-host disease or leukemia, but it was difficult to evaluate the graft-versus-leukemia effect due to the sensitivity of A20 cells to cyclophosphamide. Although the current settings provide narrow optimal therapeutic windows, further studies are warranted to maximize the benefits of each immunosuppressant.

Perivascular macrophages produce type I collagen around cerebral small vessels under prolonged hypertension in rats.

Hypertension leads to structural remodeling of cerebral blood vessels, which has been implicated in the pathophysiology of cerebrovascular diseases. The remodeling and progression of arteriolosclerosis under hypertension involve fibrosis along with the production of type I collagen around cerebral arterioles. However, the source and regulatory mechanisms of this collagen production remain elusive. In this study, we examined if perivascular macrophages (PVMs) are involved in collagen production around cerebral small vessels in hypertensive SHRSP/Izm rats. Immunoreactivity for type I collagen around cerebral small vessels in 12-week-old hypertensive rats tended to higher than those in 4-week-old hypertensive and 12-week-old control rats. In ultrastructural analyses using transmission electron microscopy, the substantial deposition of collagen fibers could be observed in the intercellular spaces around PVMs near the arterioles of rats with prolonged hypertension. In situ hybridization analyses revealed that cells positive for mRNA of Col1a1, which comprises type I collagen, were observed near cerebral small vessels. The Col1a1-positive cells around cerebral small vessels were colocalized with immunoreactivity for CD206, a marker for PVMs, but not with those for glial fibrillary acidic protein or desmin, markers for other perivascular cells such as astrocytes and vascular smooth muscle cells. These results demonstrated that enhanced production of type I collagen is observed around cerebral small vessels in rats with prolonged hypertension and Col1a1 is expressed by PVMs, and support the concept that PVMs are involved in collagen production and vascular fibrosis under hypertensive conditions.

Cluster of differentiation (CD) 9-positive mouse pituitary cells are adult stem/progenitor cells.

SOX2-positive cells are stem/progenitor cells that supply hormone-producing cells; they are found in the anterior lobe of the rodent pituitary gland. However, they are likely composed of several subpopulations. In rats, a SOX2-positive cell populations can be distinguished by the presence of S100ß. We identified the novel markers cluster of differentiation (CD) CD9 and CD81, members of the tetraspanin superfamily, for the identification of S100ß/SOX2-positive cells. Recently, CD9/CD81 double-knockout mice were generated. Although they grew normally until 3 weeks after birth, they exhibited atrophy of the pituitary gland. These findings suggested that CD9/CD81/S100ß/SOX2-positive cells in the mouse pituitary are adult stem/progenitor cells. To substantiate this hypothesis, we examined CD9 and CD81 expression in the adult and developing anterior lobe. Immunohistochemistry showed that CD9/CD81-positive cells began appearing from postnatal day 0 and settled in the stem cell niches (marginal cell layer and parenchyma) of the adult anterior lobe while expressing S100ß. We next isolated CD9 -positive cells from the adult anterior lobe, using the anti-CD9 antibody for cell characterisation. The cells in culture formed free-floating three-dimensional clusters (pituispheres); moreover, induction into all types of hormone-producing cells was successful. Furthermore, reduction of CD9 and CD81 mRNAs by siRNAs inhibited cell proliferation. These findings indicate that CD9/CD81/S100ß/SOX2-positive cells may play a role as adult stem/progenitor cells in SOX2-positive subpopulations, thus supplying hormone-producing cells in the postnatal anterior lobe. Furthermore, CD9 and CD81 are implicated in cell proliferation. The current findings provide novel insights into adult pituitary stem/progenitor cells.

ß-Cell-Specific Deletion of HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) Reductase Causes Overt Diabetes due to Reduction of ß-Cell Mass and Impaired Insulin Secretion.

Inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), statins, which are used to prevent cardiovascular diseases, are associated with a modest increase in the risk of new-onset diabetes. To investigate the role of HMGCR in the development of ß-cells and glucose homeostasis, we deleted Hmgcr in a ß-cell-specific manner by using the Cre-loxP technique. Mice lacking Hmgcr in ß-cells (ß-KO) exhibited hypoinsulinemic hyperglycemia as early as postnatal day 9 (P9) due to decreases in both ß-cell mass and insulin secretion. Ki67-positive cells were reduced in ß-KO mice at P9; thus, ß-cell mass reduction was caused by proliferation disorder immediately after birth. The mRNA expression of neurogenin3 (Ngn3), which is transiently expressed in endocrine progenitors of the embryonic pancreas, was maintained despite a striking reduction in the expression of ß-cell-associated genes, such as insulin, pancreatic and duodenal homeobox 1 (Pdx1), and MAF BZIP transcription factor A (Mafa) in the islets from ß-KO mice. Histological analyses revealed dysmorphic islets with markedly reduced numbers of ß-cells, some of which were also positive for glucagon. In conclusion, HMGCR plays critical roles not only in insulin secretion but also in the development of ß-cells in mice.

The novel ciliogenesis regulator DYRK2 governs Hedgehog signaling during mouse embryogenesis.

Mammalian Hedgehog (Hh) signaling plays key roles in embryogenesis and uniquely requires primary cilia. Functional analyses of several ciliogenesis-related genes led to the discovery of the developmental diseases known as ciliopathies. Hence, identification of mammalian factors that regulate ciliogenesis can provide insight into the molecular mechanisms of embryogenesis and ciliopathy. Here, we demonstrate that DYRK2 acts as a novel mammalian ciliogenesis-related protein kinase. Loss of Dyrk2 in mice causes suppression of Hh signaling and results in skeletal abnormalities during in vivo embryogenesis. Deletion of Dyrk2 induces abnormal ciliary morphology and trafficking of Hh pathway components. Mechanistically, transcriptome analyses demonstrate down-regulation of Aurka and other disassembly genes following Dyrk2 deletion. Taken together, the present study demonstrates for the first time that DYRK2 controls ciliogenesis and is necessary for Hh signaling during mammalian development.

Expression and functions of cluster of differentiation 9 and 81 in rat mammary epithelial cells.

Cluster of differentiation (CD) 9 and CD81 are closely-related members of the tetraspanin family that consist of four-transmembrane domain proteins. Cd9 and Cd81 are highly expressed in breast cancer cells; however, their expression in healthy mammary glands is unclear. In this study, we performed quantitative real-time PCR to analyze the expression levels of Cd9 and Cd81. Histological techniques were employed to identify Cd9- and Cd81-expressing cells in rat mammary glands during pregnancy and lactation. It was observed that Cd9 and Cd81 were expressed in the mammary glands, and their expression levels correlated with mammary gland development. To identify cells expressing Cd9 and Cd81 in the mammary glands, we performed double immunohistochemical staining for CD9 and CD81, prolactin receptor long form, estrogen receptor alpha, or Ki67. The results showed that CD9 and CD81 were co-expressed in proliferating mammary epithelial cells. Next, we attempted to isolate CD9-positive epithelial cells from the mammary gland using pluriBead cell-separation technology based on antibody-mediated binding of cells to beads of different sizes, followed by isolation using sieves with different mesh sizes. We successfully isolated CD9-positive epithelial cells with 96.8% purity. In addition, we observed that small-interfering RNAs against Cd9 and Cd81 inhibited estrogen-induced proliferation of CD9-positive mammary epithelial cells. Our current findings may provide novel insights into the proliferation of mammary epithelial cells during pregnancy and lactation as well as in pathological processes associated with breast cancer.

Differential Localization and Invasion of Tumor Cells in Mouse Models of Human and Murine Leukemias.

Leukemias are refractory hematopoietic malignancies, for which the development of new therapeutic agents requires in vivo studies using tumor-bearing mouse models. Although several organs are commonly examined in such studies to evaluate the disease course, the effectiveness of interventions and the localization of tumor cells in the affected organs are still unclear. In this study, we histologically examined the distribution of leukemia cells in several organs using two leukemic mouse models produced by the administration of two cell lines (THP-1, a human myelomonocytic leukemia, and A20, a mouse B cell leukemia/lymphoma) to severe immunodeficient mice. Survival of the mice depended on the tumor burden. Although A20 and THP-1 tumor cells massively infiltrated the parenchyma of the liver and spleen at 21 days after transplantation, A20 cells were hardly found in connective tissues in Glisson's capsule in the liver as compared with THP-1 cells. In the bone marrow, there was more severe infiltration of A20 cells than THP-1 cells. THP-1 and A20 cells were widely spread in the lungs, but were rarely observed in the small intestine. These findings suggest that each leukemia model has a unique localization of tumor cells in several affected organs, which could critically affect the disease course and the efficacy of therapeutic agents, including cellular immunotherapies.

Aldolase C is a novel molecular marker for folliculo-stellate cells in rodent pituitary.

The anterior pituitary gland is composed of five types of hormone-producing cells and folliculo-stellate cells. Folliculo-stellate cells do not produce anterior pituitary hormones but they are thought to play important roles as stem cells, phagocytes, or supporting cells of hormone-producing cells in the anterior pituitary. S100ß protein has been used as a folliculo-stellate cell marker in some animals, including rats. However, since no reliable molecular marker for folliculo-stellate cells has been reported in mice, genetic approaches for the investigation of folliculo-stellate cells in mice are not yet available. Aldolase C/Zebrin II is a brain-type isozyme and is a fructose-1,6-bisphosphate aldolase. In the present study, we first used immunohistochemistry to verify that aldolase C was produced in the anterior pituitary of rats. Moreover, using transgenic rats expressing green fluorescent protein under the control of the S100ß gene promoter, we identified aldolase C-immunoreactive signals in folliculo-stellate cells and marginal cells located in the parenchyma of the anterior pituitary and around Rathke's cleft, respectively. We also identified aldolase C-expressing cells in the mouse pituitary using immunohistochemistry and in situ hybridization. Aldolase C was not produced in any pituitary hormone-producing cells, while aldolase C-immunopositive signal co-localized with E-cadherin- and SOX2-positive cells. Using post-embedding immunoelectron microscopy, aldolase C-immunoreactive products were observed in the cytoplasm of marginal cells and folliculo-stellate cells of the mouse pituitary. Taken together, aldolase C is a common folliculo-stellate cell marker in the anterior pituitary gland of rodents.