Synergy of botanical drug extracts from Dracaena cochinchinensis stemwood and Ardisia elliptica fruit in multifunctional effects on neuroprotection and anti-inflammation.

Combination therapy is one of the promising approaches in developing therapeutics to cure complex diseases, such as Alzheimer's disease (AD). In Thai traditional medicines, the clinical application often comprises multiple botanical drugs as a formulation. The synergistic interactions between botanical drugs in combination therapies are proposed to have several advantages, including increased therapeutic efficacy, and decreased toxicity and/or adverse effects. This study aimed to explore the therapeutic functions of a botanical hybrid preparation (BHP) of two botanical drugs within a traditional multi-herbal formulation. The synergistic actions of BHP of Dracaena cochinchinensis stemwood (DCS) and Ardisia elliptica fruit (AEF) at a specific ratio of 1:9 w/w were illustrated in neuroprotection and anti-inflammation. In cultured PC12 cells, BHP of DCS and AEF showed synergistic functions in inducing neuronal differentiation, characterized by neurofilament expression and neurite outgrowth. In addition, BHP of DCS and AEF exhibited a synergistic effect in inhibiting the aggregation of Aß, a hallmark of AD pathology. The activated BV2 microglial cells induced by LPS were synergistically suppressed by the BHP of DCS and AEF, as evaluated by the expression of pro-inflammatory markers, including TNF-α, IL-1ß, and iNOS, as well as the morphological change of microglial cells. The findings suggested that the effects of BHP of DCS and AEF were greater than individual botanical drugs in a specific ratio of 1:9 w/w to enhance neuroprotective and anti-inflammatory functions.

The extracts of Dracaena cochinchinensis stemwood suppress inflammatory response and phagocytosis in lipopolysaccharide-activated microglial cells.

BACKGROUND: Neuroinflammation is a pivotal process in the brain that contributes to the development of neurodegenerative diseases, such as Alzheimer's disease (AD). During neuroinflammation, the over-activation of microglial cells can drive the pathological processes underlying AD, including an increase in amyloid ß (Aß) production and accumulation, ultimately leading to neuronal and synaptic loss. Dracaena cochinchinensis (Lour.) S.C. Chen, also known as "Chan-daeng" in Thai, belongs to the Asparagaceae family. In Thai traditional medicine, it has been used as an antipyretic, pain reliever, and anti-inflammatory agent. However, the effects of D. cochinchinensis on neuroinflammation are yet to be determined. PURPOSE: We aimed to evaluate the anti-neuroinflammatory activities of D. cochinchinensis stemwood extract in activated microglia. METHODS: In this study, lipopolysaccharide (LPS), a potent pro-inflammatory stimulus, was used to activate microglial BV2 cells, as a cell model of neuroinflammation. Our investigation included several techniques, including qRT-PCR, ELISA, Western blotting, phagocytosis, and immunofluorescence staining, to examine the potential anti-inflammatory effects of D. cochinchinensis stemwood. RESULTS: D. cochinchinensis stemwood, named DCS, was extracted with ethanol and water. The extracts of DCS showed dose-dependent anti-inflammatory effects, markedly suppressing the LPS-mediated mRNA expression of pro-inflammatory factors, including IL-1ß, TNF-α, and iNOS, while increasing expression of the anti-inflammatory biomarker Arg1 in both BV2 microglia and RAW264.7 macrophages. DCS extracts also decreased the protein levels of IL-1ß, TNF-α, and iNOS. These findings were correlated with the suppression of phosphorylated proteins of p38, JNK, and Akt in the LPS-activated microglia. Moreover, DCS extracts significantly attenuated excessive phagocytosis of beads and Aß fibrils during the LPS-mediated microglial activation. CONCLUSION: Taken together, our results indicated that DCS extracts had anti-neuroinflammatory properties by suppressing the expression of pro-inflammatory factors, increasing the expression of the anti-inflammatory biomarker Arg1, and modulating excessive phagocytosis in activated microglia. These findings suggested that DCS extract could be a promising natural product for the treatment of neuroinflammatory and neurodegenerative diseases, like AD.

Morphological profiling by high-throughput single-cell biophysical fractometry.

Complex and irregular cell architecture is known to statistically exhibit fractal geometry, i.e., a pattern resembles a smaller part of itself. Although fractal variations in cells are proven to be closely associated with the disease-related phenotypes that are otherwise obscured in the standard cell-based assays, fractal analysis with single-cell precision remains largely unexplored. To close this gap, here we develop an image-based approach that quantifies a multitude of single-cell biophysical fractal-related properties at subcellular resolution. Taking together with its high-throughput single-cell imaging performance (~10,000 cells/sec), this technique, termed single-cell biophysical fractometry, offers sufficient statistical power for delineating the cellular heterogeneity, in the context of lung-cancer cell subtype classification, drug response assays and cell-cycle progression tracking. Further correlative fractal analysis shows that single-cell biophysical fractometry can enrich the standard morphological profiling depth and spearhead systematic fractal analysis of how cell morphology encodes cellular health and pathological conditions.

Edible bird's nest, an Asian health food supplement, possesses anti-inflammatory responses in restoring the symptoms of atopic dermatitis: An analysis of signaling cascades.

Edible bird's nest (EBN) is a Chinese delicacy possessing skin rejuvenating functions. To verify skin anti-inflammatory function of EBN, water extract and enzymatic digest of EBN, as well as the major sialic acid, N-acetyl neuraminic acid (NANA), were probed in TNF-α-treated HaCaT keratinocytes. The mRNA expressions of pro-inflammatory cytokines, e.g., IL-1ß, IL-6, TNF-α, and an enzyme responsible for inflammatory response, i.e., Cox-2, as well as filaggrin and filaggrin-2, were markedly altered after treating with different preparations of EBN. The EBN-mediated responses could be accounted by its robust reduction of reactive oxygen species (ROS), NF-κB signaling and phosphorylation of p38 MAPK and JNK, as triggered by TNF-α-induced inflammation. The anti-inflammatory response of EBN was further supported in animal model. In 2,4-dinitrochlorobenzene (DNCB)-induced dermatitic mice, the effects on skin thickness, severity level of damage and scratching behavior, exerted by DNCB, were reversed after EBN treatments, in dose-dependent manners. In parallel, the levels of immune cells and pro-inflammatory cytokines in dermatitic skin were markedly reduced by treatment of EBN preparations. In general, NANA and enzymatic digest of EBN showed better anti-inflammatory responses in both models of in vitro and in vivo. These lines of evidence therefore suggest the possible application of EBN in treating atopic dermatitis.

Edible Bird's Nest, an Asian Health Food Supplement, Possesses Moisturizing Effect by Regulating Expression of Filaggrin in Skin Keratinocyte.

Edible bird's nest (EBN) has been consumed as a Chinese delicacy for hundreds of years; the functions of which have been proposed to prevent lung disease, strengthen immune response, and restore skin youthfulness. To support the skin function of EBN, the water extract and the enzymatic digest of EBN with enriched digested peptides were tested in cultured keratinocyte, HaCaT cell line. The effects of EBN extract and digest in inducing proteins crucial for skin moisturizing were determined in both in vitro and ex vivo models. In cultured keratinocytes, the expressions of S100-fused type proteins contributing to skin barrier function in the stratum corneum, e.g. filaggrin and filaggrin-2, were determined in both mRNA and protein levels, which were markedly induced in the treatment of EBN extract or digest. The EBN-induced gene transcriptions of filaggrin and filaggrin-2 were mediated by activation of p38 MAPK pathway and various transcription factors, e.g. GATA3, PPARα, PPARß, and PPARγ: these transcriptional factors were markedly activated by the digested products of EBN, as compared to the extract, in cultured keratinocytes. By using atomic force microscopy (AFM), the EBN-treated keratinocyte was shown to have more liquid-like morphology, as compared to a control cell. The EBN digest showed better induction on these moisturizing effects as compared to the extract. These lines of evidence therefore suggested the water moisturizing effect of EBN in skin function.

High-speed laser-scanning biological microscopy using FACED.

Laser scanning is used in advanced biological microscopy to deliver superior imaging contrast, resolution and sensitivity. However, it is challenging to scale up the scanning speed required for interrogating a large and heterogeneous population of biological specimens or capturing highly dynamic biological processes at high spatiotemporal resolution. Bypassing the speed limitation of traditional mechanical methods, free-space angular-chirp-enhanced delay (FACED) is an all-optical, passive and reconfigurable laser-scanning approach that has been successfully applied in different microscopy modalities at an ultrafast line-scan rate of 1-80 MHz. Optimal FACED imaging performance requires optimized experimental design and implementation to enable specific high-speed applications. In this protocol, we aim to disseminate information allowing FACED to be applied to a broader range of imaging modalities. We provide (i) a comprehensive guide and design specifications for the FACED hardware; (ii) step-by-step optical implementations of the FACED module including the key custom components; and (iii) the overall image acquisition and reconstruction pipeline. We illustrate two practical imaging configurations: multimodal FACED imaging flow cytometry (bright-field, fluorescence and second-harmonic generation) and kHz 2D two-photon fluorescence microscopy. Users with basic experience in optical microscope operation and software engineering should be able to complete the setup of the FACED imaging hardware and software in ~2-3 months.

A prepared platelet-rich plasma extract, namely Self-Growth Colony, inhibits melanogenesis by down-regulating microphthalmia-associated transcription factor in skin melanocyte.

BACKGROUND: During melanogenesis, melanocytes produce melanin through enzymatic reactions. Microphthalmia-associated transcription factor (MITF) is a major regulator in controlling the expressions of melanogenic enzymes tyrosinase (TYR), tyrosine-related protein-1 (TRP1), and dopachrome tautomerase (DCT). Self-Growth Colony (SGC) is prepared from human platelet-rich plasma (PRP) having an enrichment of growth factors, and which has claimed skin regeneration function. AIM: In this study, we aim to identify and investigate the novel role of SGC in skin melanogenesis. METHODS: MTT assay was performed to determine the cytotoxicity of applied SGC. Melanin assay was adopted to quantify the intracellular melanin after SGC treatment. Promoter-driven luciferase assay, real-time PCR, and Western blotting were performed to determine the expressions of melanogenic enzymes and MITF in SGC-treated cultured Melan-A cells, being treated with or without UV induction. Ex vivo mouse skin was treated with SGC, and then was subjected to Western blotting and histochemical staining. RESULTS: We identified that SGC inhibited melanogenesis in cultured melanocytes and ex vivo mouse skin. The phenomena were attributed to a reduction of MITF expression, which subsequently down-regulated the melanogenic enzymes, that is, TYR, TRP1, and DCT. Moreover, ERK signaling was activated in the SGC-inhibited melanogenesis. CONCLUSIONS: The findings suggest that SGC extracting from human blood can be a safe and potential agent in promoting skin whitening.

An Optimized Extract, Named Self-Growth Colony, from Platelet-Rich Plasma Shows Robust Skin Rejuvenation and Anti-Ageing Properties: A Novel Technology in Development of Cosmetics.

INTRODUCTION: Inspired by application of platelet-rich plasma (PRP) in skin treatment during injuries, an extracting method was developed here to recover high amounts of cytokines and growth factors from PRP; this prepared extract was named as self-growth colony (SGC). METHODS: In optimization of SGC preparation, various parameters were tested, for example, centrifugation force, freeze-thaw, sonication, and inclusion of calcium chelator. The amounts of cytokines and growth factors, including platelet factor 4, ß-thromboglobulin, epidermal growth factor, vascular endothelial growth factor, platelet-derived growth factor, were measured by ELISA assay. RESULTS: By comparing to PRP, the prepared SGC contained a significant higher amount of measured growth factors. In addition, the degradation of growth factors within SGC during the storage was calibrated, which showed better stability as compared to that of PRP preparation. Having possible application in skin care, the optimized SGC was chemically standardized by using the enrichment of growth factors. Application of SGC in cultured keratinocytes stimulated the wound healing of injured cultures. In line to this notion, SGC was applied onto human skin, and thereafter the robust improvement of skin properties was revealed. CONCLUSIONS: The potential application of SGC in treating skin rejuvenation and ageing, as well as its elaborated application for medical purpose, that is, wound healing, was illustrated.

Solar light induces expression of acetylcholinesterase in skin keratinocytes: Signalling mediated by activator protein 1 transcription factor.

Acetylcholinesterase (AChE) hydrolyses acetylcholine to choline and acetate, playing an important role in terminating the neurotransmission in brain and muscle. Recently, the non-neuronal functions of AChE have been proposed in different tissues, in which there are various factors to regulate the expression of AChE. In mammalian skin, AChE was identified in melanocytes and keratinocytes. Our previous study has indicated that AChE in keratinocyte affects the process of solar light-induced skin pigmentation; however, the expression of AChE in keratinocytes in responding to sunlight remains unknown. Here, we provided several lines of evidence to support a notion that AChE could be upregulated at transcriptional and translational levels in keratinocytes when exposed to solar light. The light-mediated AChE expression was triggered by Ca2+, supported by an induction of Ca2+ ionophore A23187 and a blockage by Ca2+ chelator BAPTA-AM. In addition, this increase on AChE transcriptional expression was eliminated by mutagenesis on the activating protein 1 (AP1) site in ACHE gene. Hence, the solar light-induced AChE expression is mediated by Ca2+ signalling through AP1 site. This finding supports the role of solar light in affecting the cholinergic system in skin cells, and which may further influence the dermatological function.

Parallelized volumetric fluorescence microscopy with a reconfigurable coded incoherent light-sheet array.

Parallelized fluorescence imaging has been a long-standing pursuit that can address the unmet need for a comprehensive three-dimensional (3D) visualization of dynamical biological processes with minimal photodamage. However, the available approaches are limited to incomplete parallelization in only two dimensions or sparse sampling in three dimensions. We hereby develop a novel fluorescence imaging approach, called coded light-sheet array microscopy (CLAM), which allows complete parallelized 3D imaging without mechanical scanning. Harnessing the concept of an "infinity mirror", CLAM generates a light-sheet array with controllable sheet density and degree of coherence. Thus, CLAM circumvents the common complications of multiple coherent light-sheet generation in terms of dedicated wavefront engineering and mechanical dithering/scanning. Moreover, the encoding of multiplexed optical sections in CLAM allows the synchronous capture of all sectioned images within the imaged volume. We demonstrate the utility of CLAM in different imaging scenarios, including a light-scattering medium, an optically cleared tissue, and microparticles in fluidic flow. CLAM can maximize the signal-to-noise ratio and the spatial duty cycle, and also provides a further reduction in photobleaching compared to the major scanning-based 3D imaging systems. The flexible implementation of CLAM regarding both hardware and software ensures compatibility with any light-sheet imaging modality and could thus be instrumental in a multitude of areas in biological research.

Combining technical and expert-by-experience knowledge in the quest for personal recovery from bipolar disorder: a qualitative study.

BACKGROUND: Knowledge construction is a form of communication in which people can work individually or collaboratively. Peer support services have been adopted by the public psychiatric and social welfare service as a regular form of intervention since 2015 in Hong Kong. Peer-based services can help people with bipolar disorder (BD) deal with the implications of the diagnosis, the way in which individuals with BD receive treatment, and the lifestyle changes that take place as a result of the diagnosis. Through a qualitative paradigm, this study aims to examine how individuals with BD use technical and expert-by-experience knowledge. METHODS: A total of 32 clients of mental health services were recruited from hospitals, Integrated Community Centers for Mental Wellness, and non-governmental organizations. They participated in semi-structured individual interviews. All interviews were recorded, transcribed verbatim, and analyzed using thematic analysis with the aid of NVivo. The findings were verified by peer researchers. RESULTS: Three main themes are presented in this article, including how clients made sense of the knowledge provided by mental health professionals and peer support workers (PSWs), critical perspectives about peer support services, and the way in which the services are more than knowledge transfer alone. Participants generally indicated that knowledge sharing revolved around three experiences: mood changes, medications, and sense of hope. Nevertheless, an empathic understanding of the clients' experience was more important than the sharing of knowledge. Some clients perceived medication as the chief means to recovery, so PSWs were not useful for them. However, PSW role models had an effect beyond mere knowledge transmission, as they could promote clients' pursuit of functional recovery goals. CONCLUSIONS: The present study has improved our understanding of knowledge sharing between clients with BD and health professionals or PSWs, which should take place in an empathic and hope-instilling manner. It has also emphasized the value of the presence of a role model who can speak convincingly with clients to facilitate recovery. The present findings can be used to improve the care of people with BD by generating important guidance with regard to enhancing the knowledge exchange between clients and health practitioners.

Ultrafast optical imaging at 2.0 µm through second-harmonic-generation-based time-stretch at 1.0 µm.

The performance of ultrafast time-stretch imaging at long wavelengths (beyond 1.5 µm) has suffered from low detection sensitivity due to the increasing loss of optical dispersive fibers. Here, we report an ultrafast optical imaging system with a line scan rate of ∼19 MHz at the 2.0-µm wavelength window by combining second-harmonic generation (SHG) with the highly sensitive time-stretch detection at 1.0 µm. In this imaging system, the sample is illuminated by the pulsed laser source at 2.0 µm in the spectrally encoding manner. After SHG, the encoded spectral signal at 2.0 µm is converted to 1.0 µm and then mapped to the time domain through a highly dispersive fiber at 1.0 µm, which provides a superior dispersion-to-loss ratio of ∼53 ps/nm/dB, ∼50 times larger than that of the standard fibers at 2.0 µm (typically ∼1.1 ps/nm/dB). These efforts make it possible for time-stretch technology not only being translated to longer wavelengths, where unique optical absorption contrast exists, but also benefitting from the high detection sensitivity at shorter wavelengths.

Microfluidic Imaging Flow Cytometry by Asymmetric-detection Time-stretch Optical Microscopy (ATOM).

Scaling the number of measurable parameters, which allows for multidimensional data analysis and thus higher-confidence statistical results, has been the main trend in the advanced development of flow cytometry. Notably, adding high-resolution imaging capabilities allows for the complex morphological analysis of cellular/sub-cellular structures. This is not possible with standard flow cytometers. However, it is valuable for advancing our knowledge of cellular functions and can benefit life science research, clinical diagnostics, and environmental monitoring. Incorporating imaging capabilities into flow cytometry compromises the assay throughput, primarily due to the limitations on speed and sensitivity in the camera technologies. To overcome this speed or throughput challenge facing imaging flow cytometry while preserving the image quality, asymmetric-detection time-stretch optical microscopy (ATOM) has been demonstrated to enable high-contrast, single-cell imaging with sub-cellular resolution, at an imaging throughput as high as 100,000 cells/s. Based on the imaging concept of conventional time-stretch imaging, which relies on all-optical image encoding and retrieval through the use of ultrafast broadband laser pulses, ATOM further advances imaging performance by enhancing the image contrast of unlabeled/unstained cells. This is achieved by accessing the phase-gradient information of the cells, which is spectrally encoded into single-shot broadband pulses. Hence, ATOM is particularly advantageous in high-throughput measurements of single-cell morphology and texture - information indicative of cell types, states, and even functions. Ultimately, this could become a powerful imaging flow cytometry platform for the biophysical phenotyping of cells, complementing the current state-of-the-art biochemical-marker-based cellular assay. This work describes a protocol to establish the key modules of an ATOM system (from optical frontend to data processing and visualization backend), as well as the workflow of imaging flow cytometry based on ATOM, using human cells and micro-algae as the examples.

High-throughput time-stretch imaging flow cytometry for multi-class classification of phytoplankton.

Time-stretch imaging has been regarded as an attractive technique for high-throughput imaging flow cytometry primarily owing to its real-time, continuous ultrafast operation. Nevertheless, two key challenges remain: (1) sufficiently high time-stretch image resolution and contrast is needed for visualizing sub-cellular complexity of single cells, and (2) the ability to unravel the heterogeneity and complexity of the highly diverse population of cells - a central problem of single-cell analysis in life sciences - is required. We here demonstrate an optofluidic time-stretch imaging flow cytometer that enables these two features, in the context of high-throughput multi-class (up to 14 classes) phytoplantkton screening and classification. Based on the comprehensive feature extraction and selection procedures, we show that the intracellular texture/morphology, which is revealed by high-resolution time-stretch imaging, plays a critical role of improving the accuracy of phytoplankton classification, as high as 94.7%, based on multi-class support vector machine (SVM). We also demonstrate that high-resolution time-stretch images, which allows exploitation of various feature domains, e.g. Fourier space, enables further sub-population identification - paving the way toward deeper learning and classification based on large-scale single-cell images. Not only applicable to biomedical diagnostic, this work is anticipated to find immediate applications in marine and biofuel research.