Dark Rearing in the Visual Critical Period Causes Structural Changes in Myelinated Axons in the Adult Mouse Visual Pathway.

An appropriate sensory experience during the early developmental period is important for brain maturation. Dark rearing during the visual critical period delays the maturation of neuronal circuits in the visual cortex. Although the formation and structural plasticity of the myelin sheaths on retinal ganglion cell axons modulate the visual function, the effects of dark rearing during the visual critical period on the structure of the retinal ganglion cell axons and their myelin sheaths are still unclear. To address this question, mice were reared in a dark box during the visual critical period and then normally reared to adulthood. We found that myelin sheaths on the retinal ganglion cell axons of dark-reared mice were thicker than those of normally reared mice in both the optic chiasm and optic nerve. Furthermore, whole-mount immunostaining with fluorescent axonal labeling and tissue clearing revealed that the myelin internodal length in dark-reared mice was shorter than that in normally reared mice in both the optic chiasm and optic nerve. These findings demonstrate that dark rearing during the visual critical period affects the morphology of myelin sheaths, shortens and thickens myelin sheaths in the visual pathway, despite the mice being reared in normal light/dark conditions after the dark rearing.

Bicuspidalization of the Native Tricuspid Aortic Valve: A Porcine in Vivo Model of Bicuspid Aortopathy.

Objective: To examine early histologic changes in the aorta exposed to bicuspid flow. Material and Methods: A porcine bicuspid aortopathy model was developed by suturing aortic cusps. Of nine pigs, eight underwent sham surgery (n=3) or bicuspidalization (n=5); one was used as an intact control. Wall shear stress (WSS) was assessed by computational fluid dynamics (CFD). Animals were exposed to normal or bicuspid flow for 48 h and were then euthanized for histologic examinations. Results: No animal died intraoperatively. One animal subjected to bicuspidalization died of respiratory failure during postoperative imaging studies. Echocardiography showed the aortic valve area decreased from 2.52±1.15 to 1.21±0.48 cm2 after bicuspidalization, CFD revealed increased maximum WSS (10.0±5.2 vs. 54.0±25.7 Pa; P=0.036) and percentage area of increased WSS (>5 Pa) in the ascending aorta (30.3%±24.1% vs. 81.3%±13.4%; P=0.015) after bicuspidalization. Hematoxylin-eosin staining and transmission electron microscopy showed subintimal edema and detached or degenerated endothelial cells following both sham surgery and bicuspidalization, regardless of WSS distribution. Conclusion: A bicuspid aortic valve appears to increase aortic WSS. The endothelial damage observed might have been related to non-pulsatile flow (cardiopulmonary bypass). Chronic experiments are needed to clarify the relationship between hemodynamic stress and development of bicuspid aortopathy.

Importance of Compliance With Guidelines for the Prevention of Varicella-Zoster Virus Reactivation in Multiple Myeloma.

BACKGROUND/AIM: The importance of compliance with National Comprehensive Cancer Network (NCCN) guidelines for preventing varicella-zoster virus reactivation (VZVr) in multiple myeloma (MM) in a clinical setting has not been well investigated. PATIENTS AND METHODS: We retrospectively studied the clinical characteristics and outcomes of 118 patients with MM treated with proteasome inhibitors. RESULTS: Thirty-nine episodes of VZVr were observed in 37 patients (VZVr group). The proportion of prophylactic antiviral prescriptions and compliance with antiviral prophylaxis based on the NCCN Clinical Practice guidelines was 76% and 30% in the VZVr group, and 88% and 74% in the non-VZVr group, respectively. Multivariate analysis showed that compliance with the NCCN guidelines was the only independent risk factor for VZVr (p=0.0017). CONCLUSION: It is important that prophylactic antivirals are prescribed for an appropriate duration of time to prevent the reactivation of VZV in compliance with existing guidelines.

Safety and Effectiveness of Ixazomib Dose-escalating Strategy in Ixazomib-Lenalidomide-Dexamethasone Treatment for Relapsed/Refractory Multiple Myeloma.

BACKGROUND/AIM: Gastrointestinal toxicity is common in patients receiving common therapy of ixazomib with lenalidomide and low-dose dexamethasone (IRd) for relapsed/refractory multiple myeloma. Here, we investigated the safety and effectiveness of ixazomib dosing schedules. PATIENTS AND METHODS: We retrospectively evaluated 17 consecutive patients treated with IRd (10 patients on ixazomib dose-escalation strategy (2.3 mg starting dose); seven patients on standard dose). RESULTS: The incidence of grade 3 or more haematological and grade 2 or more non-haematological adverse events was lower in the dose-escalation group than in the standard-dose group, and only that of diarrhoea was significantly lower. The median time to treatment interruption was significantly longer in the dose-escalation group than in the standard-dose group. There was no significant difference in the overall response rate (20% vs. 43%) and disease control rate (70% vs. 86%). CONCLUSION: A dose-escalation strategy to optimise ixazomib dosing may reduce treatment interruption due to adverse events without compromising its antitumor activity.

Analysis of inline-filter blockage with trastuzumab formulation using scanning-electron microscopy.

The instability of pharmaceutical monoclonal antibodies are affected by physical stimuli including, temperature, denaturant, surfactant, stirring, solid phase adsorption, oxidation, and ultraviolet rays. Clinically, we frequently experience precipitation during preparation of several pharmaceutical monoclonal antibodies for cancer. Although it is possible to remove precipitates in the injection solution during the preparation procedure, potential filter blockade during administration remains a problem with adverse effects on the quality of life of patients. Therefore we sought to investigate factors contributing to this phenomenon. To closely observed the mechanisms involved in blockade of filters during trastuzumab preparation, we prepared samples under the same conditions used in clinical practice and observed them comprehensively. The precipitates that caused filter blockade were observed when the samples were vigorously shaken and left for several hours after dissolving. The precipitates were identified as proteins. The vigorous shaking caused contact between the protein and air, which induced protein precipitation caused by the surfactants derived from the foam formation. We discovered that the external stimulation may cause the instability of monoclonal antibody preparations and, so, it is important for procedures to be as rapid to avoid precipitate formation as much as possible.

[Comparison of Chemical Behavior of Original and Generic Docetaxel Formulations as Non-alcoholic Preparations: Discussion about Diluent Solvents for Docetaxel].

　Although generic anti-tumor agents are in wide clinical use, they have not in all cases been shown to be equivalent to the original agents after preparation. In the present study, original and generic docetaxel formulations were compared with respect to stability when prepared as a non-alcoholic solution for use. When the original formulation was diluted with physiological saline solution to make a non-alcoholic preparation, the concentration decreased with time, whereas no such decrease occurred when a preparation of the generic formulation was made in a similar manner. With both the original and generic formulations, no decrease in docetaxel concentration with time was found after dilution with 5% glucose solution. On the basis of these results, it is concluded that the behaviors of original and generic docetaxel formulations are not equivalent when prepared, but that the original and generic formulations can be taken to be equivalent if they are diluted with 5% glucose solution at preparation.

Identification of M2 macrophages in anterior pituitary glands of normal rats and rats with estrogen-induced prolactinoma.

Macrophages are present throughout the anterior pituitary gland. However, the features and function of macrophages in the gland are poorly understood. Recent studies have indicated that there are two main macrophage classes: M1 (classically activated) and M2 (alternatively activated). In this study, we examine whether both M1 and M2 macrophages are present in the anterior pituitary gland of rats. Our findings indicate that macrophages that are positive for CD68 (a pan-macrophage marker) were localized near capillaries in rat anterior pituitary gland. These macrophages were positive for iNOS or mannose receptor (MR), which are markers of M1 and M2 macrophages, respectively. To determine the morphological characteristics of M2 macrophages under pathological conditions, diethylstilbestrol (DES)-treated rats were used as an animal model of prolactinoma. After 2 weeks of DES treatment, a number of MR-immunopositive cells were present in the gland. Immunoelectron microscopy revealed that MR-immunopositive M2 macrophages had many small vesicles and moderately large vacuoles in cytoplasm. Phagosomes were sometimes present in cytoplasm. Interestingly, M2 macrophages in prolactinoma tissues did not usually exhibit distinct changes or differences during the normal, hyperplasia and adenoma stages. This study is the first to confirm that both M1 and M2 macrophages are present in the anterior pituitary gland of rats. Moreover, the number of M2 macrophages was greatly increased in rats with DES-induced prolactinoma. Future studies should attempt to characterize the functional role of M2 macrophages in the gland.

Construction of a model for predicting creatinine clearance in Japanese patients treated with Cisplatin therapy.

BACKGROUND/AIM: There exist various useful predictive models, such as the Cockcroft-Gault model, for estimating creatinine clearance (CLcr). However, the prediction of renal function is difficult in patients with cancer treated with cisplatin. Therefore, we attempted to construct a new model for predicting CLcr in such patients. PATIENTS AND METHODS: Japanese patients with head and neck cancer who had received cisplatin-based chemotherapy were used as subjects. A multiple regression equation was constructed as a model for predicting CLcr values based on background and laboratory data. RESULTS: A model for predicting CLcr, which included body surface area, serum creatinine and albumin, was constructed. The model exhibited good performance prior to cisplatin therapy. In addition, it performed better than previously reported models after cisplatin therapy. CONCLUSION: The predictive model constructed in the present study displayed excellent potential and was useful for estimating the renal function of patients treated with cisplatin therapy.

In Situ Hybridization Method Reveals (Pro)renin Receptor Expressing Cells in the Pituitary Gland of Rats: Correlation with Anterior Pituitary Hormones.

Expression of (pro)renin receptor ((P)RR), a specific receptor for renin and prorenin, was studied in rat pituitary gland. In situ hybridization showed that cells expressing (P)RR mRNA were widely distributed in the anterior lobe and intermediate lobe of the pituitary gland. Double-staining using in situ hybridization for (P)RR mRNA and immunohistochemistry for the pituitary hormones showed that (P)RR mRNA was expressed in most of the GH cells and ACTH cells in the anterior lobe. (P)RR mRNA was also expressed in a few prolactin cells and TSH cells, but not in LH cells. The present study has shown for the first time the distribution of (P)RR mRNA expressing cells in the rat pituitary gland. These findings suggest that (P)RR plays physiological roles in the pituitary gland, such as the modulation of the pituitary hormone secretion.

Immunohistochemical localization of anterior pituitary hormones in S-100 protein-positive cells in the rat pituitary gland.

In the anterior and intermediate lobes of the rat pituitary gland, non-hormone-producing cells that express S-100 protein coexist with various types of hormone-producing cells and are believed to function as phagocytes, supporting and paracrine-controlling cells of hormone-producing cells and stem cells, among other functions; however, their cytological characteristics are not yet fully understood. Using a transgenic rat that expresses green fluorescent protein under the promoter of the S100ß protein gene, we immunohistochemically detected expression of the luteinizing hormone, thyroid-stimulating hormone, prolactin, growth hormone and proopiomelanocortin by S-100 protein-positive cells located between clusters of hormone-producing cells in the intermediate lobe. These findings lend support to the hypothesis that S-100 protein-positive cells are capable of differentiating into hormone-producing cells in the adult rat pituitary gland.

Spatio-temporal relation between cadherin switching and cytogenesis of hormone-producing cells in the developing rat adenohypophysis.

Cadherins are a family of transmembrane glycoproteins that mediate cell-to-cell adhesion in solid tissues and have been reported to regulate not only morphogenesis but also cell motility, proliferation, and function by activating intracellular signaling pathways. We recently found that primordial cells in the developing rat adenohypophysis co-expressed E- and N-cadherins, but endocrine cells lost E-cadherin to possess only N-cadherin at certain embryonic stages. In the present study, we aimed to elucidate the temporal relationships between cadherin expression and cell proliferation as well as between cadherin expression and the onset of hormone production in embryonic adenohypophyses. Adenohypophyses and their primordia from embryonic and postnatal rats were fixed in Bouin's fluid and paraffin sections were routinely prepared. Multiple fluorescence immunohistochemistry was performed for combinations of E-cadherin, N-cadherin, proliferating cell nuclear antigen (a marker of proliferating cells), cyclin D1, and pituitary hormones. In primordia from embryonic days 13 through 16, proliferative activities were seen in cells that co-expressed E- and N-cadherin. Cells arrested proliferation coincidentally when they lost E-cadherin after embryonic day 16. Possession of E-cadherin was closely related with expression of cyclin D1 at this stage. Moreover, hormone production was observed from embryonic day 16 only in cells that lost E-cadherin. In the developing adenohypophysis, proliferation and differentiation of hormone-producing cells have been reported to be regulated by a variety of external humoral factors. Our results raise the possibility that changes in cadherins are closely involved in these processes.

Immunohistochemical observation of co-expression of E- and N-cadherins in rat organogenesis.

Cadherins are a family of transmembrane glycoproteins that mediate cell-to-cell adhesion. Isoforms, including E- and N-cadherin, have been identified and shown to regulate morphogenesis through homophilic binding. In the ontogeny, the expressions of E- and N-cadherin change spatiotemporally, and the changes in cadherin isoforms, called cadherin switching, impact the mechanical adhesion of cells. Furthermore, cadherin functions as a receptor that transfers information from outside to inside cells, and in terms of switching, it affects cell phenotypes. To observe the expression patterns of E- and N-cadherins during embryogenesis and to identify cells that transiently coexpress both cadherins, we employed a recently developed immunohistochemical double staining technique in rat fetuses. At embryonic day 9, embryonic ectodermal cells more dominantly expressed E-cadherin, while mesodermal cells more dominantly expressed N-cadherin. At embryonic day 10, the expression pattern of E-cadherin in the surface ectoderm and endoderm and that of N-cadherin in the neuroectoderm were established. After embryonic day 10, unique co-expression of E- and N-cadherin was observed in primordia, such as the bulbus cordis, otic pit, notochord, and Rathke's pouch. In the present study, it was possible to visualize the expression patterns of E- and N-cadherin during early fetal development, which enabled us to morphologically clarify cadherin switching.

Reduction of retinaldehyde dehydrogenase 1 expression and production in estrogen-induced prolactinoma of rat.

Retinoic acid (RA) plays a critical role in normal development and tissue maintenance and is also a regulatory factor of anterior pituitary cells. We previously demonstrated that a retinoic acid-synthesizing enzyme, retinaldehyde dehydrogenase 1 (RALDH1), is expressed in prolactin cells of adult rats and that estrogen suppressed RALDH1 expression. It is suggested that RA plays a role as a paracrine and/or autocrine signaling molecule in the anterior pituitary gland. However, the presence of RALDH1 in pituitary tumors has not been demonstrated. In this study, we examined the expression of RALDH1 in diethylstilbestrol-induced prolactinoma of LEXF RI rats. Quantitative analysis of mRNA levels by real-time PCR demonstrated drastic reduction of RALDH1 expression in the prolactinoma. We have also detected both mRNA expression and production by in situ hybridization and immunohistochemistry. Both mRNA-expressing cells and immunopositive cells remarkably decreased after 4 weeks of treatment with diethylstilbestrol. Fluorescence double immunohistochemistry of RALDH1 and prolactin revealed that prolactin-immunopositive cells do not colocalize with RALDH1 in the prolactinoma. These results suggest that the reduction of local RA generation relates to cell proliferation and tumorigenesis of lactotrophs.

Changes in E- and N-cadherin expression in developing rat adenohypophysis.

Recently, we showed that hormone-producing cells express N-cadherin, while folliculo-stellate cells and marginal layer cells express E-cadherin in the adult rat anterior pituitary gland. These cells are believed to originate from a single cell population of the adenohypophyseal placode. In the present study, we immunohistochemically examined the divergence of cadherin types during pituitary histogenesis. Pituitary glands were excised from rats of embryonic day 11 (E11) through postnatal day 60 (P60) and paraffin sections were prepared. E- and N-cadherins were immunostained sequentially using monoclonal and polyclonal primary antibodies and fluorescent secondary antibodies. At E11, E-cadherin was expressed over oral epithelium, while N-cadherin expression was limited to the primordium of Rathke's pouch. When Rathke's pouch was formed at E13, E- and N-cadherin were broadly expressed in the entire cell population. N-cadherin was expressed particularly intensely in the layer of cells that faced the lumen. From E14 through E16, the majority of cells expressed both types of cadherins; however, the cell population to become the pars tuberalis expressed N-cadherin but not E-cadherin. From E18 through E20, when many hormone-producing cells appear, the number of cells that expressed N-cadherin alone increased. However, some cell populations in the pars distalis and multilayered marginal cells still expressed both cadherins. After birth, most of the cells came to express only one of the cadherin types. These results may suggest that undetermined adenohypophyseal cells express both E- and N-cadherin, but come to express either E- or N-cadherin during cytogenesis.

Distinctive localization of N- and E-cadherins in rat anterior pituitary gland.

In the rat anterior pituitary gland, folliculo-stellate cells aggregate preferably to form pseudofollicles, and each type of hormone-producing cell shows adhesive affinity with particular types of heterologous hormone-producing cells. Distribution of cadherin types in the rat anterior pituitary was examined immunohistochemically to clarify the unique cell arrangements caused by homologous and heterologous affinities among cells. N- and E-cadherins were detected continuously along cell membranes, while P-cadherin was not. N- and E-cadherins showed distinct isolation in localization, with N-cadherins localized in hormone-producing cells of distal and intermediate lobes in various amounts, and E-cadherins limited to folliculo-stellate cells and marginal layer cells facing the residual lumen of Rathke's pouch. A similar distribution of cadherins was observed in cell clusters of primary cultured anterior pituitary cells. These findings suggest that differential expression of cell adhesion molecules may be partially responsible for localization of hormone-producing cells and folliculo-stellate cells.