Nonlinear optical microscopy for skin in vivo:Basics,development and applications

Multi-photon microscopy(MPM)and coherent anti-Stokes Raman scattering(CARS)are two advanced nonlinear optical imaging techniques,which provide complementary information and have great potential in combination for noninvasive in vrito biomedical applications.This paper provides a detailed discussion of the basics,development and applications of these technologies for in vrivo skin research,covering the following topics:The principle and advantage of MPM and CARS,instrumentation development for in vino applications,MPM and CARS of normal skin,application of MPM and CARS in skin cancer and disease diagnosis;application of MPM in skin disease intervention,ie.,imaging guided two-photon photothermolysis.

Increased Cthrcl Activates Normal Fibroblasts and Suppresses Keloid Fibroblasts by Inhibiting TGF-β/Smad Signal Pathway and Modulating YAP Subcellular Location

Keloid may induce severe impairment of life quality for the patients,although keloid is a cutaneous benign tumor.Collagen triple helix repeat containing protein 1 (Cthrc1) was identified as a novel gene that was originally found in adventitial fibroblasts after arterial injury.To address the role of Cthrcl in keloid,the expression level of Cthrcl was assessed in normal skin and keloid tissue,as well as in normal fibroblasts (NFs)and keloid fibroblasts (KFs)by using quantitative PCR,Western blotting and immunohistochemical analysis.The results showed that Cthrcl was increased in keloid tissue and KFs as compared with normal skin and NFs.Meanwhile,CCK8 and Transwell assays found the cellular proliferation and migration of KFs were increased as compared with NFs.Further,to verify the function of Cthrcl in NFs and K.Fs,we increased Cthrcl expression by transfecting lentivirns (LV) vectors LV-Cthrcl.The cellular proliferation and migration,collagen synthesis and the influence on TGF-β and YAP signaling were tested.The cellular proliferation and migration were increased in NFs-Cthrcl as compared with NFs-control.Meanwhile,YAP expression and nuclear-location was increased in NFs-Cthrcl.On the contrary,when Cthrcl was overexpressed in KFs,the cellular migration was suppressed and YAP expression was reduced and transferred to cytoplasm in KFs-Cthrcl as compared with KFs-control.But the expression level of collagen I was decreased and pSmad2/3 nucleus transfer was suppressed in both NFs-Cthrc1 and KFs-Cthrc1 by using Western blotting and immunofluorescence.Increased Cthrcl activated NFs by promoting YAP nucleus translocation,whereas suppressed KFs by inhibiting YAP nucleus translocation.Enhanced Cthrcl decreased collagen I in both NFs and KFs by inhibiting TGF-β/Smad pathway.In conclusion,Cthrcl may play a role in the pathogenesis of keloid by inhibiting collagen synthesis and fibroblasts migration via suppressing TGF-β/Smad pathway and YAP nucleus translocation.

RAMAN SPECTROSCOPY FOR IN VIVO TISSUE ANALYSIS AND DIAGNOSIS,FROM INSTRUMENT DEVELOPMENT TO CLINICAL APPLICATIONS

Raman spectroscopy is a noninvasive,nondestructive analytical method capable of determining the biochemical constituents based on molecular vibrations.It does not require sample preparation or pretreatment.However,the use of Raman spectroscopy for in vivo clinical applications will depend on the feasibility of measuring Raman spectra in a relatively short time period(a few seconds).In this work,a fast dispersive-type nearinfrared(NIR)Raman spectroscopy system and a skin Raman probe were developed to facilitate real-time,noninvasive,in vivo human skin measurements.Spectrograph image aberration was corrected by a parabolic-line fiber array,permitting complete CCD vertical binning,thereby yielding a 16-fold improvement in signal-to-noise ratio.Good quality in vivo skin NIR Raman spectra free of interference from fiber fluorescence and silica Raman scattering can be acquired within one second,which greatly facilitates practical noninvasive tissue characterization and clinical diagnosis.Currently,we are conducting a large clinical study of various skin diseases in order to develop Raman spectroscopy into a useful tool for non-invasive skin cancer detection.Intermediate data analysis results are presented.Recently,we have also successfully developed a technically more challenging endoscopic Laser-Raman probe for early lung cancer detection.Preliminary in vivo results from endoscopic lung Raman measurements are discussed.

Silencing Endothelin-3 Expression Attenuates the Malignant Behaviors of Human Melanoma Cells by Regulating SPARC Levels

Summary： Endothelin-3 （ET-3） is aberrantly expressed in both metastatic melanoma tissues and cul tured melanoma cells. Our previous work showed that ET-3 could promote survival of metastatic mela noma cells via its altered expression. In this study, we investigated the mechanisms responsible for these gene-induced phenotypes in melanoma cells. An ET-3 gene sequence-specific shRNA vector pLVTHM-ET3-RNAi was constructed and transfected into human malignant melanoma cells A375 and MMRU, and the resultant molecular events and cellular changes were examined. As compared with the empty-vector group, cell proliferation was slowed down, and the growth inhibition rates were 38.9% in A375 cells and 38.4% in MMRU cells after transfection. In addition, cell invasion capability was also inhibited, with a reduction of 62.2% in A375 cells and 54.3% in MMRU cells. The percentage of apoptotic cells was found to increase. Meanwhile, in both cell lines, secreted protein acidic and rich in cy teine （SPARC） levels were down-regulated together with inhibition of its upstream signaling molecule, NF-kB. Thus, the current results suggested that down-regulated expression of ET3 attenuates the ma lignant behaviors of human melanoma ceils partially by decreasing the expression of SPARC and NF-kB.

实时拉曼光谱分析技术及其在临床早期癌症检测中的应用

拉曼光谱分析技术可以在分子水平上研究物质分子结构和生化组成信息,具有快速、准确、无创(或低创)等优点,已成为临床早期癌症检测和组织病理生理分析的重要工具。近年来,激光技术、光纤探测器件和光电检测技术的发展,不仅极大促进了新型拉曼光谱分析仪器与技术的研发,更进一步扩展了其临床应用的广度和深度,彰显出其独特的科学内涵与应用价值。对临床拉曼光谱分析技术的理论基础进行了阐述,归纳总结了临床快速拉曼光谱分析集成系统设计思路。在此基础上,以作者相关研究工作为例,探讨了拉曼光谱分析技术在临床癌症早期检测与病理分析中的应用特点,为推动相关基础研究及技术创新提供有益参考。

Childhood Langerhans cell histiocytosis:a disease with many faces

Background Langerhans cell histiocytosis(LCH)is a group of diseases characterized by the proliferation and accumulation of Langerhans cells.Clinical presentations of LCH vary widely.Data sources A PubMed search was conducted using Clinical Queries with the key term "Langerhans cell histiocytosis".The search strategy included meta-analyses,randomized controlled trials,clinical trials,observational studies,and reviews.This paper is based on,but not limited to,the search results.Results Generally,patients with LCH can be divided into two groups based on the extent of involvement at diagnosis,namely,single-system LCH and multisystem LCH.The involvement may be unifocal or multifocal.Patients with isolated bone lesions typically present between 5 and 15 years of age,whereas those with multisystem LCH tend to present before 5 years of age.The clinical spectrum is broad,ranging from an asymptomatic isolated skin or bone lesion to a life-threatening multisystem condition.Clinical manifestations include,among others,"punched out" lytic bone lesion,seborrheic dermatitis-like erup-tion,erythematous/reddish-brown crusted/scaly papules/maculopapules/plaques/patches,and eczematous lesions,diabetes insipidus,hepatosplenomegaly,cytopenias,lymphadenopathy,and an acute fulminant disseminated multisystem condition presenting with fever,skin rash,anemia,thrombocytopenia,lymphadenopathy,and hepatosplenomegaly.The diagnosis is clinicopathologic,based on typical clinical findings and histologic/immunohistochemical examination of a biopsy of lesional tissue.Positive CD1a,S100,and/or CD207(Langerin)immunohistochemical staining of lesional cells is required for a definitive diagnosis.Watchful waiting is recommended for patients with skin-only LCH.Patients with symptomatic or refractory skin-only LCH may be treated with topical tacrolimus/corticosteroids,topical nitrogen mustard,oral methotrexate,or oral hydroxyurea.The current recommended first-line therapy for patients with multisystem LCH is 12 months therapy with prednisone and vinblastine.Mercaptopurine is added for patients with risk organ involvements.Conclusions Because of the broad spectrum of clinical manifestations and the extreme diversity of disease,LCH remains a diagnostic dilemma.Morphological identification of LCH cells and positive immunochemical staining with CD1a,S100,and/or CD207(Langerin)of lesional cells are necessary for a definitive diagnosis.

Erythema nodosum

Background Erythema nodosum can be associated with a number of systemic diseases. There is, however, a paucity of information in the pediatric literature on this condition. The purpose of this article is to familiarize pediatricians with the evaluation, diagnosis, and treatment of erythema nodosum. Data sources A PubMed search was completed in Clinical Queries using the key terms 'erythema nodosum'. Results Clinically, erythema nodosum presents with a sudden onset of painful, erythematous, subcutaneous nodules mainly localized to the pretibial areas. Lesions are usually bilateral and symmetrical, ranging from 1 to 5 cm in diameter. Erythema nodosum may be associated with a variety of conditions such as infection, medications, sarcoidosis, pregnancy, inflamma-tory bowel disease, vaccination, autoimmune disease, malignancy, and miscellaneous causes. The condition is idiopathic in approximately 50% of cases. The diagnosis is mainly clinical with biopsy reserved for atypical cases. To evaluate for the underlying cause, some basic laboratory screening studies are worthwhile in most cases and include a complete blood cell count, erythrocyte sedimentation rate and/or C-reactive protein, throat swab culture, antistreptococcal O titers, and a chest radiograph. Other tests should be individualized, guided by the history and physical examination results. Most cases of ery-thema nodosum are self-limited and require no treatment. Bed rest and leg elevation are generally recommended to reduce the discomfort. Nonsteroidal anti-inflammatory drugs are the first-line treatment for pain management. Conclusions As erythema nodosum is often a cutaneous manifestation of a systemic disease, a thorough search should be performed to reveal the underlying cause.