Since mammalian skin expresses the enzymatic apparatus for melatonin synthesis, it may be an extrapineal site of melatonin synthesis. However, evidence is still lacking that this is really the case in situ. Here, we demonstrate melatonin-like immunoreactivity (IR) in the outer root sheath (ORS) of mouse and human hair follicles (HFs), which corresponds to melatonin, as shown by radioimmunoassay and liquid chromatography/tandem mass spectrometry (LC/MS/MS). The melatonin concentration in organ-cultured mouse skin, mouse vibrissae follicles, and human scalp HFs far exceeds the respective melatonin serum level and is significantly increased ex vivo by stimulation with norepinephrine (NE), the key stimulus for pineal melatonin synthesis. By real-time PCR, transcripts for the melatonin membrane receptor MT2 and for the nuclear mediator of melatonin signaling, retinoid orphan receptor alpha (ROR)alpha, are detectable in murine back skin. Transcript levels for these receptors fluctuate in a hair cycle-dependent manner, and are maximal during apoptosis-driven HF regression (catagen). Melatonin may play a role in hair cycle regulation, since its receptors (MT2 and RORalpha) are expressed in murine skin in a hair cycle-dependent manner, and because it inhibits keratinocyte apoptosis and down-regulates ERalpha expression. Therefore, the HF is both, a prominent extrapineal melatonin source, and an important peripheral melatonin target tissue. Regulated intrafollicular melatonin synthesis and signaling may play a previously unrecognized role in the endogenous controls of hair growth, for example, by modulating keratinocyte apoptosis during catagen and by desensitizing the HF to estrogen signaling. As a prototypic neuroectodermal-mesodermal interaction model, the HF can be exploited for dissecting the obscure role of melatonin in such interactions in peripheral tissues.

Diabetic kidney disease is a major cause of renal failure that urgently necessitates a breakthrough in disease management. Here we show using untargeted metabolomics that levels of phenyl sulfate, a gut microbiota-derived metabolite, increase with the progression of diabetes in rats overexpressing human uremic toxin transporter SLCO4C1 in the kidney, and are decreased in rats with limited proteinuria. In experimental models of diabetes, phenyl sulfate administration induces albuminuria and podocyte damage. In a diabetic patient cohort, phenyl sulfate levels significantly correlate with basal and predicted 2-year progression of albuminuria in patients with microalbuminuria. Inhibition of tyrosine phenol-lyase, a bacterial enzyme responsible for the synthesis of phenol from dietary tyrosine before it is metabolized into phenyl sulfate in the liver, reduces albuminuria in diabetic mice. Together, our results suggest that phenyl sulfate contributes to albuminuria and could be used as a disease marker and future therapeutic target in diabetic kidney disease.

Osteopontin (OPN) is a secreted glycoprotein with mineral- and cell-binding properties that can regulate cell activities through integrin receptors. Previously, we identified an intracellular form of osteopontin with a perimembranous distribution in migrating fetal fibroblasts (Zohar et al., J Cell Physiol 170:88-98, 1997). Since OPN and CD44 expression are increased in migrating cells, we analyzed the relationship of these proteins with immunofluorescence and confocal microscopy. A distinct co-localization of perimembranous OPN and cell-surface CD44 was observed in fetal fibroblasts, periodontal ligament cells, activated macrophages, and metastatic breast cancer cells. The co-localization of OPN and CD44 was prominent at the leading edge of migrating fibroblasts, where OPN also co-localized with the ezrin/radixin/moesin (ERM) protein ezrin, as well as in cell processes and at attachment sites of hyaluronan-coated beads. The subcortical location of OPN in these cells was verified by cell-surface biotinylation experiments in which biotinylated CD44 and non-biotinylated OPN were isolated from complexes formed with hyaluronan-coated beads and identified with immunoblotting. That perimembranous OPN represents secreted protein internalized by endocytosis or phagocytosis appeared to be unlikely since exogenous OPN that was added to cell cultures could not be detected inside the cells. A physical association with OPN, CD44, and ERM, but not with vinculin or alpha-actin, was indicated by immunoadsorption and immunoblotting of cell proteins in complexes extracted from hyaluronan-coated beads. The functional significance of OPN in this complex was demonstrated using OPN-/- and CD-/- mouse fibroblasts which displayed impaired migration and a reduced attachment to hyaluronan-coated beads. These studies indicate that OPN exists as an integral component of a hyaluronan-CD44-ERM attachment complex that is involved in the migration of embryonic fibroblasts, activated macrophages, and metastatic cells.

In this ex-vivo study, the elution components such as calcium ions from MTA had higher proliferation ability of HDP cells than control and Dycal.

These data suggest that butyrate induces apoptosis and autophagic cell death.