Role of Organic Anion Transporter NPT4 in Renal Handling of Uremic Toxin 3-indoxyl Sulfate.

Sodium-phosphate transporter NPT4 (SLC17A3) is a membrane transporter for organic anionic compounds localized on the apical membranes of kidney proximal tubular epithelial cells and plays a role in the urinary excretion of organic anionic compounds. However, its physiological role has not been sufficiently elucidated because its substrate specificity is yet to be determined. The present study aimed to comprehensively explore the physiological substrates of NPT4 in newly developed Slc17a3-/- mice using a metabolomic approach. Metabolomic analysis showed that the plasma concentrations of 11 biological substances, including 3-indoxyl sulfate, were more than two-fold higher in Slc17a3-/- mice than in wild-type mice. Moreover, urinary excretion of 3-indoxyl sulfate was reduced in Slc17a3-/- mice compared to that in wild-type mice. The uptake of 3-indoxyl sulfate by NPT4-expressing Xenopus oocytes was significantly higher than that by water-injected oocytes. The calculated Km and Vmax values for NPT4-mediated 3-indoxyl sulfate uptake were 4.52 ± 1.18 mM and 1.45 ± 0.14 nmol/oocyte/90 min, respectively. In conclusion, the present study revealed that 3-indoxyl sulfate is a novel substrate of NPT4 based on the metabolomic analysis of Slc17a3-/- mice, suggesting that NPT4 regulates systemic exposure to 3-indoxyl sulfate by regulating its urinary excretion.

Quantification of Unencapsulated Drug in Target Tissues Demonstrates Pharmacological Properties and Therapeutic Effects of Liposomal Topotecan (FF-10850).

PURPOSE: Quantifying unencapsulated drug concentrations in tissues is crucial for understanding the mechanisms underlying the efficacy and safety of liposomal drugs; however, the methodology for this has not been fully established. Herein, we aimed to investigate the enhanced therapeutic potential of a pegylated liposomal formulation of topotecan (FF-10850) by analyzing the concentrations of the unencapsulated drug in target tissues, to guide the improvement of its dosing regimen. METHODS: We developed a method for measuring unencapsulated topotecan concentrations in tumor and bone marrow interstitial fluid (BM-ISF) and applied this method to pharmacokinetic assessments. The ratios of the area under the concentration-time curves (AUCs) between tumor and BM-ISF were calculated for total and unencapsulated topotecan. DNA damage and antitumor effects of FF-10850 or non-liposomal topotecan (TPT) were evaluated in an ES-2 mice xenograft model. RESULTS: FF-10850 exhibited a much larger AUC ratio between tumor and BM-ISF for unencapsulated topotecan (2.96), but not for total topotecan (0.752), than TPT (0.833). FF-10850 promoted milder DNA damage in the bone marrow than TPT; however, FF-10850 and TPT elicited comparable DNA damage in the tumor. These findings highlight the greater tumor exposure to unencapsulated topotecan and lower bone marrow exposure to FF-10850 than TPT. The dosing regimen was successfully improved based on the kinetics of unencapsulated topotecan and DNA damage. CONCLUSIONS: Tissue pharmacokinetics of unencapsulated topotecan elucidated the favorable pharmacological properties of FF-10850. Evaluation of tissue exposure to an unencapsulated drug with appropriate pharmacodynamic markers can be valuable in optimizing liposomal drugs and dosing regimens.

Dynamic, IPSC-derived hepatic tissue tri-culture system for the evaluation of liver physiologyin vitro.

Availability of hepatic tissue for the investigation of metabolic processes is severely limited. While primary hepatocytes or animal models are widely used in pharmacological applications, a change in methodology towards more sustainable and ethical assays is highly desirable. Stem cell derived hepatic cells are generally regarded as a viable alternative for the above model systems, if current limitations in functionality and maturation can be overcome. By combining microfluidic organ-on-a-chip technology with individually differentiated, multicellular hepatic tissue fractions, we aim to improve overall functionality of hepatocyte-like cells, as well as evaluate cellular composition and interactions with non-parenchymal cell populations towards the formation of mature liver tissue. Utilizing a multi-omic approach, we show the improved maturation profiles of hepatocyte-like cells maintained in a dynamic microenvironment compared to standard tissue culture setups without continuous perfusion. In order to evaluate the resulting tissue, we employ single cell sequencing to distinguish formed subpopulations and spatial localization. While cellular input was strictly defined based on established differentiation protocols of parenchyma, endothelial and stellate cell fractions, resulting hepatic tissue was shown to comprise a complex mixture of epithelial and non-parenchymal fractions with specific local enrichment of phenotypes along the microchannel. Following this approach, we show the importance of passive, paracrine developmental processes in tissue formation. Using such complex tissue models is a crucial first step to develop stem cell-derivedin vitrosystems that can compare functionally with currently used pharmacological and toxicological applications.

Ergothioneine promotes longevity and healthy aging in male mice.

Healthy aging has emerged as a crucial issue with the increase in the geriatric population worldwide. Food-derived sulfur-containing amino acid ergothioneine (ERGO) is a potential dietary supplement, which exhibits various beneficial effects in experimental animals although the preventive effects of ERGO on aging and/or age-related impairments such as frailty and cognitive impairment are unclear. We investigated the effects of daily oral supplementation of ERGO dissolved in drinking water on lifespan, frailty, and cognitive impairment in male mice from 7 weeks of age to the end of their lives. Ingestion of 4 ~ 5 mg/kg/day of ERGO remarkably extended the lifespan of male mice. The longevity effect of ERGO was further supported by increase in life and non-frailty spans of Caenorhabditis elegans in the presence of ERGO. Compared with the control group, the ERGO group showed significantly lower age-related declines in weight, fat mass, and average and maximum movement velocities at 88 weeks of age. This was compatible with dramatical suppression by ERGO of the age-related increments in plasma biomarkers (BMs) such as the chemokine ligand 9, creatinine, symmetric dimethylarginine, urea, asymmetric dimethylarginine, quinolinic acid, and kynurenine. The oral intake of ERGO also rescued age-related impairments in learning and memory ability, which might be associated with suppression of the age-related decline in hippocampal neurogenesis and TDP43 protein aggregation and promotion of microglial shift to the M2 phenotype by ERGO ingestion. Ingestion of ERGO may promote longevity and healthy aging in male mice, possibly through multiple biological mechanisms.

Gut-liver microphysiological systems revealed potential crosstalk mechanism modulating drug metabolism.

The small intestine and liver play important role in determining oral drug's fate. Both organs are also interconnected through enterohepatic circulation, which imply there are crosstalk through circulating factors such as signaling molecules or metabolites that may affect drug metabolism. Coculture of hepatocytes and intestinal cells have shown to increase hepatic drug metabolism, yet its crosstalk mechanism is still unclear. In this study, we aim to elucidate such crosstalk by coculturing primary human hepatocytes harvested from chimeric mouse (PXB-cells) and iPSc-derived intestinal cells in a microphysiological systems (MPS). Perfusion and direct oxygenation from the MPS were chosen and confirmed to be suitable features that enhanced PXB-cells albumin secretion, cytochrome P450 (CYP) enzymes activity while also maintaining barrier integrity of iPSc-derived intestine cells. Results from RNA-sequencing showed significant upregulation in gene ontology terms related to fatty acids metabolism in PXB-cells. One of such fatty acids, arachidonic acid, enhanced several CYP enzyme activity in similar manner as coculture. From the current evidences, it is speculated that the release of bile acids from PXB-cells acted as stimuli for iPSc-derived intestine cells to release lipoprotein which was ultimately taken by PXB-cells and enhanced CYP activity.

TrkB phosphorylation in serum extracellular vesicles correlates with cognitive function enhanced by ergothioneine in humans.

Oral administration of the food-derived antioxidant amino acid ergothioneine (ERGO) results in its efficient distribution in the brain and enhances cognitive function. However, effect of ERGO deficiency on cognitive impairment and the underlying mechanisms remain unknown. We revealed that cognitive function and hippocampal neurogenesis were lower in mice fed an ERGO-free diet than in those fed the control diet. Furthermore, ERGO supplementation to achieve the control diet ERGO levels reversed these effects and restored ERGO concentrations in the plasma and hippocampus. The ERGO-induced recovery of cognitive function and hippocampal neurogenesis was blocked by inhibiting the neurotrophic factor receptor tropomyosin receptor kinase B (TrkB), with a concomitant reduction in hippocampal phosphorylated TrkB, suggesting the involvement of TrkB in these events in mice. Phosphorylated TrkB was also detected in extracellular vesicles (EVs) derived from serum of volunteers who had been orally administered placebo or ERGO-containing tablets. Importantly, the ratio of serum EV-derived phosphorylated TrkB was significantly higher in the ERGO-treated group than in the placebo-treated group and was positively correlated with both serum ERGO concentrations and several cognitive domain scores from Cognitrax. Altogether, TrkB phosphorylation is involved in ERGO-induced cognitive enhancement in mice, and TrkB phosphorylation levels in serum EVs may quantitatively represent ERGO-induced cognitive enhancement in humans.

Development of the anterior pituitary: diverse lineages of the stem/progenitor cells.

The pituitary gland is endocrine tissue composed of two distinct parts with different origins: the adenohypophysis (adenohypophyseal placode origin) and the neurohypophysis (neuroectoderm origin). Differentiation of endocrine cells in the pituitary gland leads to hormone synthesis, secretion into the capillary network, and transportation to target organs. In 1988, the discovery of the pituitary transcription factor PIT1 sparked research on endocrine cell differentiation. In the twenty-first century, the discovery that SOX2-positive stem/progenitor cells give rise to all types of pituitary endocrine cells advanced research on differentiation processes using diverse marker molecules. Lineage tracing using specific marker genes from early embryos revealed that during construction of the anterior pituitary from the adenohypophyseal placodal cells the developing anterior pituitary incorporates diverse cell types originating from the neural crest-derived and ectodermal-derived cells. Consequently, the postnatal anterior pituitary becomes a mosaic of terminally differentiated cells of different origin and with different life histories. It has also been revealed that most of the postnatal stem/progenitor cells form at least solid clusters in the parenchyma. Moreover, the classification and role of S100ß-positive cells had been ambiguous, but now they are identified as a major component of postnatal stem/progenitor cells. This paper provides an updated overview of pituitary development.

Mechanobiological stimulation in organ-on-a-chip systems reduces hepatic drug metabolic capacity in favor of regenerative specialization.

Hepatic physiology depends on the liver's complex structural composition which among others, provides high oxygen supply rates, locally differential oxygen tension, endothelial paracrine signaling, as well as residual hemodynamic shear stress to resident hepatocytes. While functional improvements were shown by implementing these factors into hepatic culture systems, direct cause-effect relationships are often not well characterized-obfuscating their individual contribution in more complex microphysiological systems. By comparing increasingly complex hepatic in vitro culture systems that gradually implement these parameters, we investigate the influence of the cellular microenvironment to overall hepatic functionality in pharmacological applications. Here, hepatocytes were modulated in terms of oxygen tension and supplementation, endothelial coculture, and exposure to fluid shear stress delineated from oxygen influx. Results from transcriptomic and metabolomic evaluation indicate that particularly oxygen supply rates are critical to enhance cellular functionality-with cellular drug metabolism remaining comparable to physiological conditions after prolonged static culture. Endothelial signaling was found to be a major contributor to differential phenotype formation known as metabolic zonation, indicated by WNT pathway activity. Lastly, oxygen-delineated shear stress was identified to direct cellular fate towards increased hepatic plasticity and regenerative phenotypes at the cost of drug metabolic functionality - in line with regenerative effects observed in vivo. With these results, we provide a systematic evaluation of critical parameters and their impact in hepatic systems. Given their adherence to physiological effects in vivo, this highlights the importance of their implementation in biomimetic devices, such as organ-on-a-chip systems. Considering recent advances in basic liver biology, direct translation of physiological structures into in vitro models is a promising strategy to expand the capabilities of pharmacological models.

A highly efficient cell culture method using oxygen-permeable PDMS-based honeycomb microwells produces functional liver organoids from human induced pluripotent stem cell-derived carboxypeptidase M liver progenitor cells.

Recent advancements in bioengineering have introduced potential alternatives to liver transplantation via the development of self-assembled liver organoids, derived from human-induced pluripotent stem cells (hiPSCs). However, the limited maturity of the tissue makes it challenging to implement this technology on a large scale in clinical settings. In this study, we developed a highly efficient method for generating functional liver organoids from hiPSC-derived carboxypeptidase M liver progenitor cells (CPM+ LPCs), using a microwell structure, and enhanced maturation through direct oxygenation in oxygen-permeable culture plates. We compared the morphology, gene expression profile, and function of the liver organoid with those of cells cultured under conventional conditions using either monolayer or spheroid culture systems. Our results revealed that liver organoids generated using polydimethylsiloxane-based honeycomb microwells significantly exhibited enhanced albumin secretion, hepatic marker expression, and cytochrome P450-mediated metabolism. Additionally, the oxygenated organoids consisted of both hepatocytes and cholangiocytes, which showed increased expression of bile transporter-related genes as well as enhanced bile transport function. Oxygen-permeable polydimethylsiloxane membranes may offer an efficient approach to generating highly mature liver organoids consisting of diverse cell populations.

The balance between fetal oxytocin and placental leucine aminopeptidase (P-LAP) controls human uterine contraction around labor onset.

A fetal pituitary hormone, oxytocin which causes uterine contractions, increases throughout gestation, and its increase reaches 10-fold from week 32 afterward. Oxytocin is, on the other hand, degraded by placental leucine aminopeptidase (P-LAP) which exists in both terminal villi and maternal blood. Maternal blood P-LAP increases with advancing gestation under the control of non-genomic effects of progesterone, which is also produced from the placenta. Progesterone is converted to estrogen by CYP17A1 localized in the fetal adrenal gland and placenta at term. The higher oxytocin concentrations in the fetus than in the mother demonstrate not only fetal oxytocin production but also its degradation and/or inhibition of leakage from fetus to mother by P-LAP. Until labor onset, the pregnant uterus is quiescent possibly due to the balance between increasing fetal oxytocin and P-LAP under control of progesterone. A close correlation exists between the feto-placental and maternal units in the placental circulation, although the blood in the two circulations does not necessarily mix. Fetal maturation results in progesterone withdrawal via the CYP17A1 activation accompanied with fetal oxytocin increase. Contribution of fetal oxytocin to labor onset has been acknowledged through the recognition that the effect of fetal oxytocin in the maternal blood is strictly regulated by its degradation by P-LAP under the control of non-genomic effects of progesterone. In all senses, the fetus necessarily takes the initiative in labor onset.

Renal Pharmacokinetic Adaptation to Cholestasis Causes Increased Nephrotoxic Drug Accumulation by Mrp6 Downregulation in Mice.

In hepatic dysfunction, renal pharmacokinetic adaptation can be observed, although information on the changes in drug exposure and the interorgan regulation of membrane transporters in kidney in liver diseases is limited. This study aimed to clarify the effects of renal exposure to nephrotoxic drugs during cholestasis induced by bile duct ligation (BDL). Among the 11 nephrotoxic drugs examined, the tissue accumulation of imatinib and cisplatin in kidney slices obtained from mice 2 weeks after BDL operation was higher than that in sham-operated mice. The uptake of imatinib in the kidney slices of BDL mice was slightly higher, whereas its efflux from the slices was largely decreased compared to that in sham-operated mice. Proteomic analysis revealed a reduction in renal expression of the efflux transporter multidrug resistance-associated protein 6 (Mrp6/Abcc6) in BDL mice, and both imatinib and cisplatin were identified as Mrp6 substrates. Survival probability after cisplatin administration was reduced in BDL mice. In conclusion, the present study demonstrated that BDL-induced cholestasis leads to the downregulation of the renal basolateral efflux transporter Mrp6, resulting in drug accumulation in renal cells and promoting drug-induced renal injury.

Missing pieces of the pituitary puzzle: participation of extra-adenohypophyseal placode-lineage cells in the adult pituitary gland.

The pituitary gland is a major endocrine tissue composing of two distinct entities, the adenohypophysis (anterior pituitary, cranial placode origin) and the neurohypophysis (posterior pituitary, neural ectoderm origin), and plays important roles in maintaining vital homeostasis. This tissue is maintained by a slow, consistent cell-renewal system of adult stem/progenitor cells. Recent accumulating evidence shows that neural crest-, head mesenchyme-, and endoderm lineage cells invade during pituitary development and contribute to the maintenance of the adult pituitary gland. Based on these novel observations, this article discusses whether these lineage cells are involved in pituitary organogenesis, maintenance, regeneration, dysplasia, or tumors.

l-Type amino acid transporter 1 in hypothalamic neurons in mice maintains energy and bone homeostasis.

Hypothalamic neurons regulate body homeostasis by sensing and integrating changes in the levels of key hormones and primary nutrients (amino acids, glucose, and lipids). However, the molecular mechanisms that enable hypothalamic neurons to detect primary nutrients remain elusive. Here, we identified l-type amino acid transporter 1 (LAT1) in hypothalamic leptin receptor-expressing (LepR-expressing) neurons as being important for systemic energy and bone homeostasis. We observed LAT1-dependent amino acid uptake in the hypothalamus, which was compromised in a mouse model of obesity and diabetes. Mice lacking LAT1 (encoded by solute carrier transporter 7a5, Slc7a5) in LepR-expressing neurons exhibited obesity-related phenotypes and higher bone mass. Slc7a5 deficiency caused sympathetic dysfunction and leptin insensitivity in LepR-expressing neurons before obesity onset. Importantly, restoring Slc7a5 expression selectively in LepR-expressing ventromedial hypothalamus neurons rescued energy and bone homeostasis in mice deficient for Slc7a5 in LepR-expressing cells. Mechanistic target of rapamycin complex-1 (mTORC1) was found to be a crucial mediator of LAT1-dependent regulation of energy and bone homeostasis. These results suggest that the LAT1/mTORC1 axis in LepR-expressing neurons controls energy and bone homeostasis by fine-tuning sympathetic outflow, thus providing in vivo evidence of the implications of amino acid sensing by hypothalamic neurons in body homeostasis.

Emerging Roles of Uremic Toxins and Inflammatory Cytokines in the Alteration of Hepatic Drug Disposition in Patients with Kidney Dysfunction.

Patients with kidney dysfunction exhibit distinct pharmacokinetic profiles compared to those with normal kidney function. Hence, it is desirable to monitor the drug efficacy and toxicity caused by fluctuations in plasma drug concentrations associated with kidney dysfunction. Recently, pharmacokinetic information of drugs excreted mainly through the urine of patients with kidney dysfunction has been reported via drug-labeling information. Pharmacokinetic changes in drugs mainly eliminated by the liver cannot be overlooked as drug metabolism and/or transport activity in the liver may also be altered in patients with kidney dysfunction; however, the underlying mechanisms remain unclear. To plan an appropriate dosage regimen, it is necessary to clarify the underlying processes of functional changes in pharmacokinetic proteins. In recent years, uremic toxins have been shown to reduce the activity and/or expression of renal and hepatic transporters. This inhibitory effect has been reported to be time-dependent. In addition, inflammatory cytokines, such as interleukin-6, released from immune cells activated by uremic toxins and/or kidney injury can reduce the expression levels of drug-metabolizing enzymes and transporters in human hepatocytes. In this mini-review, we have summarized the renal and hepatic pharmacokinetic changes as well as the potential underlying mechanisms in kidney dysfunction, such as the chronic kidney disease and acute kidney injury. SIGNIFICANCE STATEMENT: Patients with kidney dysfunction exhibit distinct pharmacokinetic profiles compared to those with normal kidney function. Increased plasma concentrations of uremic toxins and inflammatory cytokines during kidney disease may potentially affect the activities and/or expression levels of drug-metabolizing enzymes and transporters in the liver and kidneys.

Adeno-associated virus-mediated knockdown demonstrates the major role of hepatic Bcrp in the overall disposition of the active metabolite of the tyrosine kinase inhibitor regorafenib in mice.

Breast cancer resistance protein (BCRP) is expressed on hepatic bile canalicular membranes; however, its impact on substrate drug disposition is limited. This study proposes an in vivo knockdown approach using adeno-associated virus encoding short hairpin RNA (shRNA) targeting the bcrp gene (AAV-shBcrp) to clarify the substrate, the overall disposition of which is largely governed by hepatic Bcrp. The disposition of the tyrosine kinase inhibitor, regorafenib, was first examined in bcrp gene knockout (Bcrp-/-) and wild-type (WT) mice, as it was sequentially converted to active metabolites M - 2 and M - 5, which are BCRP substrates. After oral administration of regorafenib, plasma and liver concentrations of M - 5, but not regorafenib, were higher in Bcrp-/- than WT mice. To directly examine the role of hepatic Bcrp in M - 5 disposition, M - 5 was intravenously injected into mice three weeks after the intravenous injection of AAV-shBcrp, when mRNA of Bcrp in the liver (but not the small intestine) was downregulated. AAV-shBcrp-treated mice showed higher M - 5 concentration in plasma and liver, but lower biliary excretion than the control mice, indicating the fundamental role of hepatic Bcrp in M - 5 disposition. This is the first application of AAV-knockdown strategy to clarify the pharmacokinetic role of xenobiotic efflux transporters in the liver.

Enhancement and maintenance of hepatic metabolic functions by controlling 3D aggregation of cryopreserved human iPS cell-derived hepatocyte-like cells.

Hepatocyte-like cells (HLCs) generated from human induced pluripotent stem cells are potent cells to study individual-specific hepatotoxicity for drug screening test. However, the functions of metabolic enzymes are practically low. Here, we reconstituted stable and compact 3D spheroids of commercially available cryopreserved HLCs by our original spheroid formation method with high viscous methylcellulose medium. 3D formation enhanced the hepatic functions and maintained the functions for 14 days. Especially, the expression of cytochrome P450s was 10- to 100-fold enhanced compared to conventional 2D culture, which is applicable to a typical drug-metabolizing test using liquid chromatograph-tandem mass spectrometer. In conclusion, we successfully formed human HLC spheroid from commercially available cryo-preserved cells, which realized remarkable hepatic maturation by prolonged 3D culture, especially in terms of drug-metabolizing enzymes. Our spheroid formation technology has the potential to make HLC spheroids a potent tool in aspects of pharmaceutical research, such as drug screening and pharmacokinetic studies.

A physiologically based pharmacokinetic and pharmacodynamic model for disposition of FF-10832.

This study aimed to quantitatively clarify the critical factors responsible for the superior antitumor efficacy of a liposomal gemcitabine (2,2-difluorodeoxycytidine; dFdC) formulation, FF-10832, compared with dFdC. The underlying hypothesis is the different exposure of tumors to its active metabolite, dFdC triphosphate (dFdCTP), between the two formulations. Therefore, physiologically based pharmacokinetic and pharmacodynamic (PBPK/PD) models for encapsulated and unencapsulated dFdC were constructed considering the tumor dFdCTP concentration as an index of antitumor activity. To estimate drug the parameters, the time profiles of encapsulated and unencapsulated dFdC in the blood and those of dFdC and dFdCTP in tumors were measured following the intravenous bolus administration of FF-10832 or dFdC. dFdC metabolism and transport in the liver S9 fraction and isolated hepatocytes, respectively, were experimentally determined. The tumor growth curve in a mouse xenograft model following the administration of FF10832 and dFdC was also used to construct the PD model. The sensitivity analysis of the PBPK/PD model revealed the critical factors affecting antitumor efficacy, which included the total and intratumor tissue uptake clearances for liposomal formulation and the cytidine deaminase and deoxycytidine deaminase activities in tumors. Thus, these parameters are potential biomarkers for predicting the efficacy of the liposomal formulation of dFdC.

Accurate Evaluation of Hepatocyte Metabolisms on a Noble Oxygen-Permeable Material With Low Sorption Characteristics.

In the pharmaceutical industry, primary cultured hepatocytes is a standard tool used to assess hepatic metabolisms and toxicity in vitro. Drawbacks, however, include their functional deterioration upon isolation, mostly due to the lack of a physiological environment. Polydimethylsiloxane (PDMS) has been reported to improve the function of isolated hepatocytes by its high oxygen permeability when used as a material of microphysiological systems (MPS). However, its high chemical sorption property has impeded its practical use in drug development. In this study, we evaluated a new culture material, 4-polymethyl-1-pentene polymer (PMP), in comparison with PDMS and conventional tissue culture polystyrene (TCPS). First, we confirmed the high oxygen permeability and low sorption of PMP, and these properties were comparable with PDMS and TCPS, respectively. Moreover, using primary rat hepatocytes, we demonstrated maintained high levels of liver function at least for 1 week on PMP, with its low chemical sorption and high oxygen permeability being key factors in both revealing the potential of primary cultured hepatocytes and in performing an accurate evaluation of hepatic metabolisms. Taken together, we conclude that PMP is a superior alternative to both PDMS and TCPS, and a promising material for a variety of drug testing systems.

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.

Crosstalk between foetal vasoactive peptide hormones and placental aminopeptidases regulates placental blood flow: Its significance in preeclampsia.

In pregnancy, placental circulation occurs through two independent circulation systems: foetoplacental and uterine (spiral artery)-placental lake. Crosstalk between the foetal peptide hormones, angiotensin II (A-II) and vasopressin (AVP), and their degrading placental aminopeptidases (APs), aminopeptidase A for A-II and placental leucine aminopeptidase for both AVP and oxytocin, primarily regulate placental circulation. On the other hand, placental circulation represents an arteriovenous shunt. In normal pregnancy, the blood pressure decreases, despite increased cardiac output and plasma volume, probably due to the arteriovenous shunt in the growing placenta. Actually, the foetal vasoactive hormones in the foetoplacental circulation are much higher than those in the maternal circulation throughout pregnancy. In normal pregnancy, AP activity derived from the placenta in maternal blood increases with gestation and placental growth. Foetal hypoxia increases the secretion of foetal both AVP and A-II. Although there is an increase in both AP activities in the maternal blood in normal pregnancy, their activities increase more than those in normal pregnancy during mild preeclampsia. However, both AP activities decline significantly compared than those in severe preeclampsia. This suggests that AP prevents leakage of increased foetal vasoactive hormones into the maternal blood in mild preeclampsia, and its protective role breaks down in severe preeclampsia, leading to a massive leak of the hormones into maternal circulation and consequent marked contraction of both the maternal vessels and the uterus. Consequently, AP activity in both placenta and maternal blood acts as the foeto-maternal barrier for foetal vasoactive hormones and thus contributes to the onset of preeclampsia.