Rapid induction of dopaminergic neuron-like cells from human fibroblasts by autophagy activation with only 2-small molecules.

The dopaminergic neurons are responsible for the release of dopamine. Several diseases that affect motor function, including Parkinson's disease (PD), are rooted in inadequate dopamine (DA) neurotransmission. The study's goal was to create a quick way to make dopaminergic neuron-like cells from human fibroblasts (hNF) using only two small molecules: hedgehog pathway inhibitor 1 (HPI-1) and neurodazine (NZ). Two small compounds have been shown to induce the transdifferentiation of hNF cells into dopaminergic neuron-like cells. After 10 days of treatment, hNF cells had a big drop in fibroblastic markers (Col1A1, KRT18, and Elastin) and a rise in neuron marker genes (TUJ1, PAX6, and SOX1). Different proteins and factors related to dopaminergic neurons (TH, TUJ1, and dopamine) were significantly increased in cells that behave like dopaminergic neurons after treatment. A study of the autophagy signaling pathway showed that apoptotic genes were downregulated while autophagy genes (LC3, ATG5, and ATG12) were significantly upregulated. Our results showed that treating hNF cells with both HPI-1 and NZ together can quickly change them into mature neurons that have dopaminergic activity. However, the current understanding of the underlying mechanisms involved in nerve guidance remains unstable and complex. Ongoing research in this field must continue to advance for a more in-depth understanding. This is crucial for the safe and highly effective clinical application of the knowledge gained to promote neural regeneration in different neurological diseases.

3D culture of alginate-hyaluronic acid hydrogel supports the stemness of human mesenchymal stem cells.

The three-dimensional (3D) cell culture system is being employed more frequently to investigate cell engineering and tissue repair due to its close mimicry of in vivo microenvironments. In this study, we developed natural biomaterials, including hyaluronic acid, alginate, and gelatin, to mimic the creation of a 3D human mesenchymal stem cell (hMSC) extracellular environment and selected hydrogels with high proliferation capacity for 3D MSC culture. Human mesenchymal stem cells were encapsulated within hydrogels, and an investigation was conducted into the effects on cell viability and proliferation, stemness properties, and telomere activity compared to the 2D monolayer culture. Hydrogel characterization, cell proliferation, Live/Dead cell viability assay, gene expression, telomere relative length, and MSC stemness-related proteins by immunofluorescence staining were examined. The results showed that 3D alginate-hyaluronic acid (AL-HA) hydrogels increased cell proliferation, and the cells were grown as cellular spheroids within hydrogels and presented a high survival rate of 77.36% during the culture period of 14 days. Furthermore, the 3D alginate-hyaluronic acid (AL-HA) hydrogels increased the expression of stemness-related genes (OCT-4, NANOG, SOX2, and SIRT1), tissue growth and development genes (YAP and TAZ), and cell proliferation gene (Ki67) after culture for 14 days. Moreover, the telomere activity of the 3D MSCs was enhanced, as indicated by the upregulation of the human telomerase reverse transcriptase gene (hTERT) and the relative telomere length (T/S ratio) compared to the 2D monolayer culture. Altogether, these data suggest that the 3D alginate-hyaluronic acid (AL-HA) hydrogels could serve as a promising material for maintaining stem cell properties and might be a suitable carrier for tissue engineering proposals.

Nanoencapsulated cordyceps extract enhances collagen synthesis and skin cell regeneration through antioxidation and autophagy.

Oxidative stress from reactive oxygen species is the main cause of skin ageing. Cordycepin, a bioactive compound of Cordyceps militaris, contains antioxidant activity. This study examined extracellular matrix, antioxidant effect, autophagy activity, and skin regeneration in human dermal fibroblasts (HDFs) under normal and oxidative stress conditions. Slow disintegration was used to create nano-encapsulated cordyceps extract. HDFs were cultured and treated with 1 M cordycepin, 1 M medium, 0.1 M cordyceps medium loaded nanoparticles (CMP), or 1 mM H2O2. HDFs' senescent phenotypes were assessed, including cell proliferation, ROS scavenging, collagen and elastin synthesis, antioxidant activity, and wound healing. CMP size averaged 184.5 ± 95.2 nm increased cell proliferation and reduced H2O2-induced ROS. Thus, HDFs treated for 48 h increased skin regeneration activity 2.76-fold by expressing extracellular matrix and rescuing H2O2-induced damaged cells. It was significant that this CMP inhibited H2O2-induced oxidative stress and induced autophagy to regenerate HDFs. The developed CMP could be used in cosmetics.

Enhancing Neurological Competence of Nanoencapsulated Cordyceps/Turmeric Extracts in Human Neuroblastoma SH-SY5Y Cells.

Introduction: Neurological diseases, including Alzheimer's, Parkinson's diseases, and brain cancers, are reportedly caused by genetic aberration and cellular malfunction. Herbs with bioactive compounds that have anti-oxidant effects such as cordyceps and turmeric, are of interest to clinical applications due to their minimal adverse effects. The aim of study is to develop the nanoencapsulated cordyceps and turmeric extracts and investigate their capability to enhance the biological activity and improve neuronal function. Methods: Human neuroblastoma SH-SY5Y cells were utilized as a neuronal model to investigate the properties of nanoencapsulated cordyceps or turmeric extracts, called CMP and TEP, respectively. SH-SY5Y cells were treated with either CMP or TEP and examined the biological consequences, including neuronal maturation and neuronal function. Results: The results showed that both CMP and TEP improved cellular uptake efficiency within 6 h by 2.3 and 2.8 times, respectively. Besides, they were able to inhibit cellular proliferation of SH-SY5Y cells up to 153- and 218-fold changes, and increase the expression of mature neuronal markers (TUJ1, PAX6, and NESTIN). Upon the treatment of CMP and TEP, the expression of dopaminergic-specific genes (LMX1B, FOXA2, EN1, and NURR1), and the secretion level of dopamine were significantly improved up to 3.3-fold and 3.0-fold, respectively, while the expression of Alzheimer genes (PSEN1, PSEN2, and APP), and the secretion of amyloid precursor protein were significantly reduced by 32-fold and 108-fold, respectively. Importantly, the autophagy activity was upregulated by CMP and TEP at 6.3- and 5.5-fold changes, respectively. Conclusions: This finding suggested that the nanoencapsulated cordyceps and turmeric extracts accelerated neuronal maturation and alleviated neuronal pathology in human neural cells. This paves the way for nanotechnology-driven drug delivery systems that could potentially be used as an alternative medicine in the future for neurological diseases.

Cordycepin-loaded Nanoparticles from Cassava Starch Promote the Proliferation of Submandibular Gland Cells and Inhibit the Growth of Oral Squamous Carcinoma Cells.

This study examined associations between the effect of treatment with nano-cassava starch that contained cordycepin (CS) extract, targeting human submandibular gland cells (HSGs), and human oral squamous carcinoma cells (HSC-4). Cassava starch nanoparticles (CSNPs) were prepared by either physical or acid treatment. These nanoparticles were then loaded with either CS or cordyceps medium and then treated with HSG or HSC-4 cells in different concentrations of CS and nanoparticles. Moreover, the protein secretion, reactive oxygen species (ROS) activity and the expression of salivary-specific genes, antioxidant gene were determined after treatment. CSNPs can enhance the activity of CS at low concentrations. Cordycepin-loaded cassava starch nanoparticles (CCSNPs) increased HSG proliferation, protein secretion, and the expression of salivary-specific genes, AMY and AQP5. Besides, CCSNPs also protected and scavenged of ROS via the stimulation of the antioxidant genes in HSGs, indicating the protective roles of CS to HSGs. On the other hand, CCSNPs inhibited the growth of HSC-4 cells by stimulating ROS generation and reducing protein secretion. This finding suggested that CCSNPs presented the dual actions against HSGs and human oral squamous carcinoma cells, and the encapsulation of CS with cassava nanoparticles enhanced the activity of CS.

Cordycepin attenuates Salivary Hypofunction through the Prevention of Oxidative Stress in Human Submandibular Gland Cells.

Xerostomia (dry mouth) is a significant age-related condition. Meanwhile, cordycepin, the natural therapeutic agent, has demonstrated an anti-aging effect. Therefore, the present study aimed to investigate the preventive effects of cordycepin on secretory function in an in vitro model of hydrogen peroxide (H2O2)-induced salivary hypofunction. After being exposed to H2O2, human submandibular gland (HSG) cells were treated with various concentrations of cordycepin (6.25-50 µM) for 24, 48, and 72h. To evaluate cell proliferation and reactive oxygen species (ROS) generation, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and 2, 7'-dichlorodihydrofluorescein diacetate assays were performed. The amylase activity was kinetically measured by 2-chloro-p-nitrophenol linked with maltotrioside. The expression of salivary, antioxidant and apoptotic markers at mRNA and protein levels were performed by reverse transcriptase polymerase chain reaction (RT-PCR) and immunofluorescence analysis, respectively. We demonstrated that cordycepin (6.25-25 µM) contributed to significant increases in expression of the salivary marker genes, alpha-amylase 1 (AMY1A) and aquaporin-5 (AQP5), and in amylase secretion without changes in cell viability. Under oxidative stress, HSG cells showed remarkable dysfunction. Cordycepin rescued the protective effects partially by decreasing ROS generation and restoring the expression of the salivary proteins, AMY and AQP5 via anti-oxidant and anti-apoptotic activity. In addition, the amount of amylase that was secreted from HSG cells cultured in cordycepin was increased. In conclusion, cordycepin demonstrated a protective effect on H2O2 -induced HSG cells by decreasing ROS generation and upregulating the salivary function markers, AMY1A and AQP5, at both the transcriptional and translational levels.