Metabolic Engineering of Escherichia coli for Enhanced Production of Naringenin 7-Sulfate and Its Biological Activities.

Flavonoids are one of the predominant groups of plant polyphenols, and these compounds have significant effects on human health and nutrition. Sulfated flavonoids have more favorable attributes compared to their parent compounds such as increased solubility, stability, and bioavailability. In this research, we developed a microbial system to produce sulfated naringenin using Escherichia coli expressing a sulfotransferase (ST) from Arabidopsis thaliana (At2g03770). This wild-type strain was used as a model system for testing clustered regularly interspaced short palindromic repeats (CRISPR) interference (CRISPRi) metabolic engineering strategies. Using synthetic sgRNA to mediate transcriptional repression of cysH, a gene encoding 3'-phosphoadenosine-5'-phosphosulfate (PAPS) ST, which is involved in sulfur metabolism, resulted in an increase in intracellular PAPS accumulation by over 3.28-fold without impairing cell growth. Moreover, naringenin 7-sulfate production by engineering E. coli with its cysH gene repressed in the open reading frame through CRISPRi was enhanced by 2.83-fold in compared with the wild-type control. To improve the efficiency of biotransformation, the concentration of SO42- , glucose, and substrate were optimized. The bioproductivity of naringenin 7-sulfate was 135.49 µM [∼143.1 mg (47.7 mg L-1)] in a 3-L fermenter at 36 h. These results demonstrated that the CRISPRi system was successfully applied for the first time in E. coli to develop an efficient microbial strain for production of a sulfated flavonoid. In addition, antibacterial and anticancer activities of naringenin 7-sulfate were investigated and found to be higher than the parent compound.

Facial skin barrier function recovery after microneedle transdermal delivery treatment.

BACKGROUND: Microneedle treatment is currently used in the cosmetic industry for several skin conditions. Despite their extensive use, there is lack of sufficient data on the safety of microneedles. OBJECTIVE: To investigate the degree of acute skin damage and the time required for facial skin barrier function to recover using different microneedle lengths and numbers of applications. MATERIALS AND METHODS: Each side of a volunteer's face was randomly treated with one of the following treatments: five applications of 0.15-mm microneedles, five applications of 0.25-mm microneedles, 10 applications of 0.15-mm microneedles, or 10 applications of 0.25-mm microneedles. Transepidermal water loss, stratum corneum hydration, and skin erythema were measured at baseline, immediately after treatment, 4 hours after treatment, and 8 hours after treatment and at 24-hour intervals for 3 days. RESULTS: Prompt recovery of barrier function (within 72 hours) was observed after microneedle treatment. CONCLUSION: Microneedle treatment is simple and inexpensive, and the skin barrier disruption it causes resolves quickly. Therefore, it can serve as an effective physical method of enhancing transdermal delivery of medications for the treatment of many cosmetic and dermatological conditions.

Suppression of tumor necrosis factor-alpha and inducible nitric oxide synthase gene expression by THI 52, a new synthetic naphthyl-benzylisoquinoline alkaloid.

The effects of THI 52 (1-naphthylethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline) on (a) inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) expression in RAW 264.7 cells stimulated by lipopolysaccharide (LPS)/interferon gamma (IFN-gamma), (b) plasma nitrate concentration as well as iNOS protein expression (lung) in vivo in LPS-treated rats, and (c) the restoration of vascular contractility to vasoconstrictor agents in LPS-treated vessels in vitro were investigated. THI 52 concentration-dependently reduced not only nitric oxide (NO) production (IC(50) value, 12.5 microM) but also the expression of TNF-alpha and iNOS mRNA in RAW 264.7 cells. Incubation of rat endothelium-denuded thoracic aorta with LPS (300 ng/mL) in vitro for 8 hr resulted in the suppression of vasoconstrictor effects to phenylephrine (PE), effects that were restored by co-incubation with THI 52. Administration of THI 52 (10 and 20mg/kg, i.p.) 30 min before injection of LPS (10mg/kg, i.p.) resulted in a significant reduction of the expression of iNOS protein in rat lung tissue and in the plasma nitrite/nitrate (NOx) level. Addition of THI 52-treated macrophage-conditioned medium to a TNF-sensitive L929 fibroblast cell line (CCL1) increased cell viability, depending on the concentration of THI 52. Finally, THI 52 inhibited the activation of nuclear factor kappaB (NF-kappaB) by inhibition of IkappaB degradation through the prevention of IkappaB phosphorylation. Collectively, these results strongly suggest that THI 52 suppresses both TNF-alpha and iNOS gene expression by inhibiting NF-kappaB. Thus, THI 52, a new synthetic isoquinoline alkaloid, may be beneficial in inflammatory disorders where the overproduction of NO and TNF-alpha is a matter of concern.