Signalling by senescent melanocytes hyperactivates hair growth.

Niche signals maintain stem cells in a prolonged quiescence or transiently activate them for proper regeneration1. Altering balanced niche signalling can lead to regenerative disorders. Melanocytic skin nevi in human often display excessive hair growth, suggesting hair stem cell hyperactivity. Here, using genetic mouse models of nevi2,3, we show that dermal clusters of senescent melanocytes drive epithelial hair stem cells to exit quiescence and change their transcriptome and composition, potently enhancing hair renewal. Nevus melanocytes activate a distinct secretome, enriched for signalling factors. Osteopontin, the leading nevus signalling factor, is both necessary and sufficient to induce hair growth. Injection of osteopontin or its genetic overexpression is sufficient to induce robust hair growth in mice, whereas germline and conditional deletions of either osteopontin or CD44, its cognate receptor on epithelial hair cells, rescue enhanced hair growth induced by dermal nevus melanocytes. Osteopontin is overexpressed in human hairy nevi, and it stimulates new growth of human hair follicles. Although broad accumulation of senescent cells, such as upon ageing or genotoxic stress, is detrimental for the regenerative capacity of tissue4, we show that signalling by senescent cell clusters can potently enhance the activity of adjacent intact stem cells and stimulate tissue renewal. This finding identifies senescent cells and their secretome as an attractive therapeutic target in regenerative disorders.

Widespread somatic L1 retrotransposition in normal colorectal epithelium.

Throughout an individual's lifetime, genomic alterations accumulate in somatic cells1-11. However, the mutational landscape induced by retrotransposition of long interspersed nuclear element-1 (L1), a widespread mobile element in the human genome12-14, is poorly understood in normal cells. Here we explored the whole-genome sequences of 899 single-cell clones established from three different cell types collected from 28 individuals. We identified 1,708 somatic L1 retrotransposition events that were enriched in colorectal epithelium and showed a positive relationship with age. Fingerprinting of source elements showed 34 retrotransposition-competent L1s. Multidimensional analysis demonstrated that (1) somatic L1 retrotranspositions occur from early embryogenesis at a substantial rate, (2) epigenetic on/off of a source element is preferentially determined in the early organogenesis stage, (3) retrotransposition-competent L1s with a lower population allele frequency have higher retrotransposition activity and (4) only a small fraction of L1 transcripts in the cytoplasm are finally retrotransposed in somatic cells. Analysis of matched cancers further suggested that somatic L1 retrotransposition rate is substantially increased during colorectal tumourigenesis. In summary, this study illustrates L1 retrotransposition-induced somatic mosaicism in normal cells and provides insights into the genomic and epigenomic regulation of transposable elements over the human lifetime.

Restoration of Immune Privilege in Human Dermal Papillae Controlling Epithelial-Mesenchymal Interactions in Hair Formation.

BACKGROUND: Hair follicles are among a handful of organs that exhibit immune privilege. Dysfunction of the hair follicle immune system underlies the development of inflammatory diseases, such as alopecia areata. METHODS: Quantitative reverse transcription PCR and immunostaining was used to confirm the expression of major histocompatibility complex class I in human dermal papilla cells. Through transcriptomic analyses of human keratinocyte stem cells, major histocompatibility complex class I was identified as differentially expressed genes. Organ culture and patch assay were performed to assess the ability of WNT3a conditioned media to rescue immune privilege. Lastly, CD8+ T cells were detected near the hair bulb in alopecia areata patients through immunohistochemistry. RESULTS: Inflammatory factors such as tumor necrosis factor alpha and interferon gamma were verified to induce the expression of major histocompatibility complex class I proteins in dermal papilla cells. Additionally, loss of immune privilege of hair follicles was rescued following treatment with conditioned media from outer root sheath cells. Transcriptomic analyses found 58 up-regulated genes and 183 down-regulated genes related in MHC class I+ cells. Using newborn hair patch assay, we demonstrated that WNT3a conditioned media with epidermal growth factor can restore hair growth. In alopecia areata patients, CD8+ T cells were increased during the transition from mid-anagen to late catagen. CONCLUSION: Identification of mechanisms governing epithelial and mesenchymal interactions of the hair follicle facilitates an improved understanding of the regulation of hair follicle immune privilege.

Bayesian log-normal deconvolution for enhanced in silico microdissection of bulk gene expression data.

Deconvolution of bulk gene expression profiles into the cellular components is pivotal to portraying tissue's complex cellular make-up, such as the tumor microenvironment. However, the inherently variable nature of gene expression requires a comprehensive statistical model and reliable prior knowledge of individual cell types that can be obtained from single-cell RNA sequencing. We introduce BLADE (Bayesian Log-normAl Deconvolution), a unified Bayesian framework to estimate both cellular composition and gene expression profiles for each cell type. Unlike previous comprehensive statistical approaches, BLADE can handle > 20 types of cells due to the efficient variational inference. Throughout an intensive evaluation with > 700 simulated and real datasets, BLADE demonstrated enhanced robustness against gene expression variability and better completeness than conventional methods, in particular, to reconstruct gene expression profiles of each cell type. In summary, BLADE is a powerful tool to unravel heterogeneous cellular activity in complex biological systems from standard bulk gene expression data.

Modified Transosseous Wiring Technique for Neglected Fracture-Dislocation of the Proximal Interphalangeal Joint.

BACKGROUND: Fracture-dislocation of the proximal interphalangeal (PIP) joint of the finger is challenging due to the high risk of stiffness. The purpose of this study is to evaluate the clinical and radiological results of a modified transosseous wiring technique for the management of chronic fracture-dislocations of the PIP joint. METHODS: Ten patients (nine men and one woman; mean age, 38.3 years; range, 21 to 69 years) with neglected fracture-dislocation of the PIP joint were included. The mean duration from injury to operation was 14.7 weeks (range, 3 to 66 weeks). The dorsolateral approach and extension block pinning were used to reduce dislocation. After thorough debridement of the scar tissues in the dorsal dead space and the fracture site, the reduction was maintained with transosseous wiring. Radiologic evaluations of bone union and arthritic changes and clinical evaluations (range of motion of the PIP joint and Disabilities of the Arm, Shoulder and Hand [DASH] score) were performed. The mean follow-up period was 12.9 months (range, 12 to 19 months). RESULTS: All patients demonstrated evidence of radiographic healing within a mean healing time of 6 weeks (range, 4 to 10 weeks); however, one had a widened gap and one had an early arthritic change. The mean range of motion in the PIP joint was 81° (range, 50° to 105°). The mean DASH score was 21.6 (range, 7.5 to 35.8). CONCLUSIONS: For chronic fracture-dislocation of the PIP joint, transosseous wiring with direct curettage and optimal bone purchase can provide satisfying outcome.

A Safer Non-surgical Filler Augmentation Rhinoplasty Based on the Anatomy of the Nose.

BACKGROUND: Filler augmentation rhinoplasty is a quick, non-surgical procedure that can produce outcomes comparable to open rhinoplasty surgery. However, the increased frequency of vascular complications has emerged as an important issue. The present study aimed to investigate measures to overcome the vascular complications based on the anatomy of the nose. METHODS: A colored filler was injected into cadavers for augmentation of the nasal dorsum using the retrograde injection technique and direct percutaneous injection technique. The concavity of the sellion area was measured using lateral view cephalography X-ray images. Lastly, we used ultrasonography to determine filler location in 20 Korean patients who had filler injected into the sellion area by injection at the infratip lobule. RESULTS: Filler was injected into the superficial layer by the retrograde injection technique in three cadavers and into the deep layer by direct percutaneous injection technique in another three cadavers. The average angle between the nasal dorsum skin and sellion was found to be 10.2 ± 2.8 degrees, while the minimum angle was 5.1 degrees. The average distance between the needle tip and nasal bone was 1.9 ± 0.3 mm, while the minimum distance was 0.4 mm. CONCLUSIONS: When performing filler augmentation rhinoplasty on the sellion area, direct percutaneous injection from the glabella can allow more accurate injection into the supraperiosteal level, which can reduce complications such as visual loss and skin necrosis due to vascular compromise. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

In Situ Synthesis of Bimetallic Tungsten-Copper Nanoparticles via Reactive Radio-Frequency (RF) Thermal Plasma.

We synthesize, in situ, W-x wt% Cu (x = 5, 10, and 20 wt%) composite nanoparticles using inductively coupled radio-frequency (RF) thermal plasma. In the RF thermal plasma process, the W-x wt% Cu composite nanoparticles are synthesized by hydrogen reduction of tungsten trioxide (WO3) and cupric oxide (CuO). The synthesized W and Cu nanoparticles are effectively reduced to W and Cu, and the W-Cu nanoparticles are uniformly distributed bimetallic (or composite) nanoparticles.

Estrogen modulates mesenchyme-epidermis interactions in the adult nipple.

Maintenance of specialized epidermis requires signals from the underlying mesenchyme; however, the specific pathways involved remain to be identified. By recombining cells from the ventral skin of the K14-PTHrP transgenic mice [which overexpress parathyroid hormone-related protein (PTHrP) in their developing epidermis and mammary glands] with those from wild type, we show that transgenic stroma is sufficient to reprogram wild-type keratinocytes into nipple-like epidermis. To identify candidate nipple-specific signaling factors, we compared gene expression signatures of sorted Pdgfrα-positive ventral K14-PTHrP and wild-type fibroblasts, identifying differentially expressed transcripts that are involved in WNT, HGF, TGFß, IGF, BMP, FGF and estrogen signaling. Considering that some of the growth factor pathways are targets for estrogen regulation, we examined the upstream role of this hormone in maintaining the nipple. Ablation of estrogen signaling through ovariectomy produced nipples with abnormally thin epidermis, and we identified TGFß as a negatively regulated target of estrogen signaling. Estrogen treatment represses Tgfß1 at the transcript and protein levels in K14-PTHrP fibroblasts in vitro, while ovariectomy increases Tgfb1 levels in K14-PTHrP ventral skin. Moreover, ectopic delivery of Tgfß1 protein into nipple connective tissue reduced epidermal proliferation. Taken together, these results show that specialized nipple epidermis is maintained by estrogen-induced repression of TGFß signaling in the local fibroblasts.

Role of 14-3-3 sigma in over-expression of P-gp by rifampin and paclitaxel stimulation through interaction with PXR.

In this study, we presented the role of 14-3-3σ to activate CK2-Hsp90ß-PXR-MDR1 pathway on rifampin and paclitaxel treated LS174T cells and in vivo LS174T cell-xenografted nude mouse model. Following several in vitro and in vivo experiments, rifampin and paclitaxel were found to be stimulated the CK2-Hsp90ß-PXR-MDR1 pathway. Of the proteins in this pathway, Pregnane X receptor (PXR) is a representative transcription factor of multidrug resistance protein 1 (MDR1). We constructed FLAG-PXR-LS174T stable cell lines and discovered 22 proteins that interacted with PXR on rifampin treatment. Among them, Hsp90ß and 14-3-3σ were isolated for further study. Both the proteins were found to be localized in cytoplasm on rifampin treatment by using confocal microscopy. On the other hand, PXR was found to be localized in nucleus after rifampin and paclitaxel treatment by using cell fractionation assay. In Western blot analysis, rifampin did not influence the expression of 14-3-3σ protein. Transient transfection of 14-3-3σ into LS174T cells induced overexpression of PXR; however, P-glycoprotein (P-gp) was not changed significantly. P-gp overexpression was induced only when 14-3-3σ transfected LS174T cells were treated with rifampin and paclitaxel, whereas 14-3-3σ inhibition by nonpeptidic inhibitor, BV02 and 14-3-3σ siRNA reduced rifampin induced PXR and P-gp expression. Cell survival rates were much higher at 14-3-3σ-LS174T stable cell lines than LS174T cells following paclitaxel and vincristine treatment. This data indicates that 14-3-3σ contributes to P-gp overexpression through interaction with PXR with rifampin and paclitaxel treatment.

Does the Gadoxetic Acid-Enhanced Liver MRI Impact on the Treatment of Patients with Colorectal Cancer? Comparison Study with ¹8F-FDG PET/CT.

OBJECTIVES: We evaluated the value of Gadoxetic acid-enhanced liver MRI in the preoperative staging of colorectal cancer and estimated the clinical impact of liver MRI in the management plan of liver metastasis. METHODS: We identified 108 patients who underwent PET/CT and liver MRI as preoperative evaluation of colorectal cancer, between January 2011 and December 2013. We evaluated the per nodule sensitivity of PET/CT and liver MRI for liver metastasis. Management plan changes were estimated for patients with metastatic nodules newly detected on liver MRI, to assess the clinical impact. RESULTS: We enrolled 131 metastatic nodules (mean size 1.6 cm) in 41 patients (mean age 65 years). The per nodule sensitivities of PET/CT and liver MRI were both 100% for nodules measuring 2 cm or larger but were significantly different for nodules measuring less than 2 cm (59.8% and 95.1%, resp., P = 0.0001). At least one more metastatic nodule was detected on MRI in 16 patients. Among these, 7 patients indicated changes of management plan after performing MRI. CONCLUSIONS: Gadoxetic acid-enhanced liver MRI detected more metastatic nodules compared with PET/CT, especially for small (<2 cm) nodules. The newly detected nodules induced management plan change in 43.8% (7/16) of patients.

A Guide to Studying Human Hair Follicle Cycling In Vivo.

Hair follicles (HFs) undergo lifelong cyclical transformations, progressing through stages of rapid growth (anagen), regression (catagen), and relative "quiescence" (telogen). Given that HF cycling abnormalities underlie many human hair growth disorders, the accurate classification of individual cycle stages within skin biopsies is clinically important and essential for hair research. For preclinical human hair research purposes, human scalp skin can be xenografted onto immunocompromised mice to study human HF cycling and manipulate long-lasting anagen in vivo. Although available for mice, a comprehensive guide on how to recognize different human hair cycle stages in vivo is lacking. In this article, we present such a guide, which uses objective, well-defined, and reproducible criteria, and integrates simple morphological indicators with advanced, (immuno)-histochemical markers. This guide also characterizes human HF cycling in xenografts and highlights the utility of this model for in vivo hair research. Detailed schematic drawings and representative micrographs provide examples of how best to identify human HF stages, even in suboptimally sectioned tissue, and practical recommendations are given for designing human-on-mouse hair cycle experiments. Thus, this guide seeks to offer a benchmark for human hair cycle stage classification, for both hair research experts and newcomers to the field.

Diffusion-weighted MRI of epithelial ovarian cancers: correlation of apparent diffusion coefficient values with histologic grade and surgical stage.

OBJECTIVE: The purpose of this article is to correlate the apparent diffusion coefficient (ADC) values of epithelial ovarian cancers with histologic grade and surgical stage. MATERIALS AND METHODS: We enrolled 43 patients with pathologically proven epithelial ovarian cancers for this retrospective study. All patients underwent preoperative pelvic magnetic resonance imaging (MRI) including diffusion-weighted images with b value of 0 and 1000 s/mm2 at 3.0-T unit. The mean ADC values of the solid portion of the tumor were measured and compared among different histologic grades and surgical stages. RESULTS: The mean ADC values of epithelial ovarian cancers differed significantly between grade 1 (well-differentiated) and grade 2 (moderately-differentiated) (P=0.013) as well as between grade 1 and grade 3 (poorly-differentiated) (P=0.01); however, no statistically significant difference existed between grade 2 and grade 3 (P=0.737). The receiver-operating characteristic analysis indicated that a cutoff ADC value of less than or equal to 1.09×10(-3)mm2/s was associated with 94.4% sensitivity and 85.7% specificity in distinguishing grade 1 and grade 2/3 cancer. The difference in mean ADC values was statistically significant for early stage (FIGO stage I) and advanced stage (FIGO stage II-IV) cancer (P=0.011). The interobserver agreement for the mean ADC values of epithelial ovarian cancers was excellent. CONCLUSION: The mean ADC values of the solid portion of epithelial ovarian cancers negatively correlated to histologic grade and surgical stage. The mean ADC values may be useful imaging biomarkers for assessment of tumor grade of epithelial ovarian cancer.

Erythropoietin promotes hair shaft growth in cultured human hair follicles and modulates hair growth in mice.

BACKGROUND: Recent studies have shown that erythropoietin (EPO)/erythropoietin receptor (EPOR) signaling exist in both human and mouse hair follicles (HFs). OBJECTIVE: To investigate whether dermal papilla cells (DPCs) express functional EPOR and, if so, to investigate effects of EPO on hair shaft growth in cultured human scalp hair follicles and hair growth in mice. METHODS: EPOR expression in DPCs and follicular keratinocytes was examined by RT-PCR and immunoblot. Phosphorylation of EPOR signaling pathway mediators by EPO treatment was examined by immunoblot. MTT assay was employed to check cell viability after EPO treatment. Hair shaft growth was measured in the absence or presence of EPO and matrix keratinocyte proliferation was examined by Ki-67 immunostaining in cultured hair follicles. Agarose beads containing EPO were implanted into dorsal skin of C57BL/6 mice to examine effects of EPO on hair growth in vivo. RESULTS: EPOR mRNA and protein are expressed in cultured human DPCs. EPOR signaling pathway mediators such as EPOR and Akt are phosphorylated by EPO in DPCs. EPO significantly promoted the growth of DPCs and elongated hair shafts with increased proliferation of matrix keratinocytes in cultured human hair follicles. In addition, EPO not only promoted anagen induction from telogen but also prolonged anagen phase. CONCLUSIONS: EPO may modulate hair growth by stimulating DPCs that express functional EPOR.

Identification of transcriptional targets of Wnt/beta-catenin signaling in dermal papilla cells of human scalp hair follicles: EP2 is a novel transcriptional target of Wnt3a.

BACKGROUND: Recent studies showed that Wnt signaling through the beta-catenin pathway (canonical Wnt signaling) act on mouse dermal papilla cells (DPCs) enabling hair follicles to keep growing. OBJECTIVE: To investigate whether human DPCs respond to canonical Wnt signaling and, if so, to identify target genes of Wnt/beta-catenin pathway. METHODS: Cultured human DPCs were transiently transfected with the beta-catenin responsive TCF reporter plasmid (pTopflash) and corresponding negative control reporter (pFopflash) to assess the activity of beta-catenin signaling by Wnt3a (one of the canonical Wnts). Immunofluorescence staining was also performed to localize beta-catenin in the presence or absence of Wnt3a. Microarray was carried out using Affymetrix gene chips. RT-PCR analysis and immunoblot were employed to verify microarray data. Cyclic AMP (cAMP) levels were measured using EIA assay after Wnt3a and PGE2 treatment in DPCs. RESULTS: Wnt3a significantly stimulated the transcriptional activity of pTopflash but not pFopflash. In line with this, we identified a number of genes that are regulated by Wnt3a. Some of the differently expressed genes including EP2 were confirmed by RT-PCR analysis. Immunoblot further confirmed that EP2 protein is indeed increased by Wnt3a. DPCs pretreated with Wnt3a showed higher responsiveness to PGE2 as measured by cAMP levels. CONCLUSIONS: Elucidation of the role of Wnt3a-regulated genes identified in this study including EP2 would help our understanding of hair-induction and maintenance of anagen phase.