Photoprotective effects of Lithospermum erythrorhizon and Pueraria lobata extracts on UVB-induced photoaging: A study on skin barrier protection.

AIM: Lithospermum erythrorhizon and Pueraria lobata exhibit promising potential as cosmetic additives for mitigating skin barrier impairment induced by photoaging. Despite their potential, the precise mechanisms underlying their protective and ameliorative effects remain elusive. This study sought to assess the reparative properties of Lithospermum erythrorhizon and Pueraria lobata extracts (LP) on UVB-irradiated human skin keratinocytes (HaCaT cells) and explore the therapeutic potential of LP as a skin barrier protection agent. MATERIALS AND METHODS: Antioxidant activities were gauged through 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and reactive oxygen species (ROS) assays. The expression levels of skin barrier-related markers, encompassing metalloproteinases (MMPs) and hyaluronidase (HYAL) were scrutinized using enzyme-linked immunosorbent assay (ELISA), reverse transcriptase (RT)-PCR, and Western blotting, with a particular focus on the involvement of the transforming growth factor (TGF)-ß/Smad and nuclear factor-κB (NF-κB) signaling pathways. RESULTS: The study revealed that LP effectively scavenges free radicals, diminishes ROS production in a dose-dependent manner, and significantly attenuates UVB-induced expression of MMP-1 and MMP-3 through modulation of the hyaluronan synthase (HAS)2/HYAL1 signaling axis in UVB-irradiated HaCaT cells. Additionally, LP demonstrated enhanced TGF-ß signaling activation, fostering procollagen type I synthesis, and concurrently exhibited mitogen-activated protein kinases (MAPK)/NF-κB signaling inactivation, thereby mitigating pro-inflammatory cytokine release and alleviating UVB-induced cellular damage. CONCLUSION: In conclusion, the observed protective effects of LP on skin cellular constituents highlight its substantial biological potential for shielding against UVB-induced skin photoaging, positioning it as a promising candidate for both pharmaceutical and cosmetic applications.

Clinical efficacy of herbal extracts in treatment of mild to moderate acne vulgaris: an 8-week, double-blinded, randomized, controlled trial.

BACKGROUND: Herbal extracts with fewer adverse effects can be an alternative to these drugs because they can target various molecular pathways of acne pathogenesis. OBJECTIVES: To evaluate the clinical efficacy of herbal extracts (mangosteen, Lithospermum officinale, Tribulus terrestris L., Houttuynia cordata Thunb) for the treatment of mild to moderate acne vulgaris. METHODS: Sixty patients were randomized in a 1:1 ratio to receive blinded treatment with herbal extracts or vehicle for 8 weeks. Inflammatory and non-inflammatory acne lesion counts, Investigator's Global Assessment, patient's satisfaction and safety profiles were assessed. We also performed skin biopsy at baseline and week 8 to confirm immunological changes with immunohistochemistry staining. RESULTS: By the end of the study period, both inflammatory and non-inflammatory acne lesion counts were significantly decreased in herbal extracts group (p< .05). In immunohistochemistry staining, expressions of IL-1α, IL-8, and keratin 16 were significantly decreased in herbal extracts group compared to vehicle group from baseline to week 8. There was no serious adverse events in both groups. CONCLUSIONS: This herbal extracts can be a new therapeutic option for patients with mild to moderate acne vulgaris who are reluctant to use drugs.

Comparative Proteomic Analysis of Lithospermum erythrorhizon Reveals Regulation of a Variety of Metabolic Enzymes Leading to Comprehensive Understanding of the Shikonin Biosynthetic Pathway.

Plants produce a large variety of specialized (secondary) metabolites having a wide range of hydrophobicity. Shikonin, a red naphthoquinone pigment, is a highly hydrophobic metabolite produced in the roots of Lithospermum erythrorhizon, a medicinal plant in the family Boraginaceae. The shikonin molecule is formed by the coupling of p-hydroxybenzoic acid and geranyl diphosphate, catalyzed by a membrane-bound geranyltransferase LePGT at the endoplasmic reticulum, followed by cyclization of the geranyl chain and oxidations; the latter half of this biosynthetic pathway, however, has not yet been clarified. To shed light on these steps, a proteome analysis was conducted. Shikonin production in vitro was specifically regulated by illumination and by the difference in media used to culture cells and hairy roots. In intact plants, however, shikonin is produced exclusively in the root bark of L. erythrorhizon. These features were utilized for comparative transcriptome and proteome analyses. As the genome sequence is not known for this medicinal plant, sequences from de novo RNA-seq data with 95,861 contigs were used as reference for proteome analysis. Because shikonin biosynthesis requires copper ions and is sensitive to blue light, this methodology identified strong candidates for enzymes involved in shikonin biosynthesis, such as polyphenol oxidase, cannabidiolic acid synthase-like and neomenthol dehydrogenase-like proteins. Because acetylshikonin is the main end product of shikonin derivatives, an O-acetyltransferase was also identified. This enzyme may be responsible for end product formation in these plant species. Taken together, these findings suggest a putative pathway for shikonin biosynthesis.

De Novo Transcriptome Assembly and Characterization of Lithospermum officinale to Discover Putative Genes Involved in Specialized Metabolites Biosynthesis.

Lithospermum officinale is a valuable source of bioactive metabolites with medicinal and industrial values. However, little is known about genes involved in the biosynthesis of these metabolites, primarily due to the lack of genome or transcriptome resources. This study presents the first effort to establish and characterize de novo transcriptome assembly resource for L. officinale and expression analysis for three of its tissues, namely leaf, stem, and root. Using over 4Gbps of RNA-sequencing datasets, we obtained de novo transcriptome assembly of L. officinale, consisting of 77,047 unigenes with assembly N50 value as 1524 bps. Based on transcriptome annotation and functional classification, 52,766 unigenes were assigned with putative genes functions, gene ontology terms, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. KEGG pathway and gene ontology enrichment analysis using highly expressed unigenes across three tissues and targeted metabolome analysis showed active secondary metabolic processes enriched specifically in the root of L. officinale. Using co-expression analysis, we also identified 20 and 48 unigenes representing different enzymes of lithospermic/chlorogenic acid and shikonin biosynthesis pathways, respectively. We further identified 15 candidate unigenes annotated as cytochrome P450 with the highest expression in the root of L. officinale as novel genes with a role in key biochemical reactions toward shikonin biosynthesis. Thus, through this study, we not only generated a high-quality genomic resource for L. officinale but also propose candidate genes to be involved in shikonin biosynthesis pathways for further functional characterization.

RNA-seq transcriptome analysis of breast cancer cell lines under shikonin treatment.

Shikonin is a naphthoquinone isolated from the dried root of Lithospermum erythrorhizon, an herb used in Chinese medicine. Although several studies have indicated that shikonin exhibits antitumor activity in breast cancer, the mechanism of action remains unclear. In the present study, we performed transcriptome analysis using RNA-seq and explored the mechanism of action of shikonin in regulating the growth of different types of breast cancer cells. The IC50 of shikonin on MCF-7, SKBR-3 and MDA-MB-231 cells were 10.3 µΜ, 15.0 µΜ, 15.0 µΜ respectively. Our results also demonstrated that shikonin arrests the progression of cell cycle and induces apoptosis in MDA-MB-231 cells. Using RNA-seq transcriptome analysis, we found 38 common genes that significantly express in different types of breast cancer cells under shikonin treatment. In particular, our results indicated that shikonin induces the expression of dual specificity phosphatase (DUSP)-1 and DUSP2 in both RNA and protein levels. In addition, shikonin also inhibits the phosphorylation of JNK and p38, the downstream signaling molecules of DUSP1 and DUSP2. Therefore, our results suggest that shikonin induces the expression of DUSP1 and DUSP2 which consequently switches off JNK and p38 MAPK pathways and causes cell cycle arrest and apoptosis in breast cancer cells.

[Production of shikonin by cell cultures of Lithospermum erythrorhizon].

OBJECTIVE: To explore cultural conditions of shikonin production by cell cultures of Lithospermum erythrorhizon. METHOD: Orthogonal design was applied in determination of shikonin within the medium. Flask test was applied in the study of shikonin production by the amount of ventilation. RESULT: The best medium consisted of 100 mg/L L-phenylalanine, 2 mg/L IAA and 800 mg/L Ca(NO3)2 4H2O. The best amount of ventilation was get by shaken at 150 r/min. CONCLUSION: This test provided data for producing shikonin by cell cultures of Lithospermum erythrorhizon.