Promoting FADH2 Regeneration of Hydroxylation for High-Level Production of Hydroxytyrosol from Glycerol in Escherichia coli.

Hydroxytyrosol is a natural polyphenolic compound widely used in the food and drug industries. The current commercial production of hydroxytyrosol relies mainly on plant extracts, which involve long extraction cycles and various raw materials. Microbial fermentation has potential value as an environmentally friendly and low-cost method. Here, a de novo biosynthetic pathway of hydroxytyrosol has been designed and constructed in an Escherichia coli strain with released tyrosine feedback inhibition. By introduction of hpaBC from E. coli and ARO10 and ADH6 from Saccharomyces cerevisiae, the de novo biosynthesis of hydroxytyrosol was achieved. An important finding in cofactor engineering is that the introduction of L-amino acid deaminase (LAAD) promotes not only cofactor regeneration but also metabolic flow redistribution. To further enhance the hydroxylation process, different 4-hydroxyphenylacetate 3-monooxygenase (hpaB) mutants and HpaBC proteins from different sources were screened. Finally, after optimization of the carbon source, pH, and seed medium, the optimum engineered strain produced 9.87 g/L hydroxytyrosol in a 5 L bioreactor. This represents the highest titer reported to date for de novo biosynthesis of hydroxytyrosol in microorganisms.

Trifluoperazine Synergistically Potentiates Bortezomib-Induced Anti-Cancer Effect in Multiple Myeloma via Inhibiting P38 MAPK/NUPR1.

Multiple myeloma (MM) is a common hematological malignancy. Bortezomib (BTZ) is a traditional medicine for MM treatment, but there are limitations for current treatment methods. Trifluoperazine (TFP) is a clinical drug for acute and chronic psychosis therapy. Lately, researchers have found that TFP can suppress tumor growth in many cancers. We attempted to study the effects of BTZ and TFP on MM in vivo and in vitro. We concentrated on the individual and combined impact of BTZ and TFP on the proliferation and apoptosis of MM cells via Cell Counting kit-8 assay, EdU assay, western blot, and flow cytometry. We found that combination therapy has a strong synergistic impact on MM cells. Combination therapy could induce cell arrest during G2/M phase and induce apoptosis in MM cells. Meanwhile, BTZ combined with TFP could play a better role in the anti-MM effect in vivo through MM.1s xenograft tumor models. Furthermore, we explored the mechanism of TFP-induced apoptosis in MM, and we noticed that TFP might induce MM apoptosis by inhibiting p-P38 MAPK/NUPR1. In summary, our findings suggest that TFP could synergistically enhance the BTZ-induced anti-cancer effect in multiple myeloma, which might be a promising therapeutic strategy for MM treatment.

Clinical observation of depression after breast cancer operation treated with aurieular point sticking therapy / 中国针灸

<p><b>OBJECTIVE</b>To compare the differences in the efficacy on depression after breast cancer operation treated with auricular point sticking therapy, the combined program of auricular point sticking therapy and TCM psychotherapy and medication with deanxit.</p><p><b>METHODS</b>Ninety patients were randomized into 3 groups, 30 cases in each one. In the western medication group, the simple oral administration of deanxit was applied, one tablet a day. In the auricular point group, Xin (GO15), Shen (CO10), Gan(CO12), Shenmen(TF4), Pizhixia (AT), Neifenmi (CO18) were selected and stimulated with auricular point sticking on either side in each treatment, once a week. In the combined program group, on the basis of the treatment as the auricular point group, TCM psychotherapy was combined with. The treatment of 4 weeks made one session. One session and 4 weeks follow-up were required. The self-rating depression scale (SDS) was used to compare the score before and after treatment in the patients of each group and the efficacy was assessed.</p><p><b>RESULTS</b>(1) Compared with those before treatment, in 4 weeks of treatment and 4 weeks of follow-up, SDS scores were all reduced apparently in the 3 groups (all P<0.001). After 4 weeks of follow-up in the auricular point group, after 2 and 4 weeks of treatment and 4 weeks of follow-up in the combined program group, SDS scores were all lower than those in the western medication group (all P<0.05). (2) After 4 weeks of treatment, the curative rate in the combined program group was higher than that in the auricular point group and the western medication group [60.0% (18/30) vs. 40.0% (12/30), 36.7% (11/30), both P<0.05)]. After 4 weeks of follow-up, the curative rate in either the combined program group or the auricular point group was higher than that in the western medication group [60.0% (18/30), 30.0% (19/30) vs. 23.3% (7/30), both P<0.05].</p><p><b>CONCLUSION</b>The auricular point sticking therapy, the combined program of auricular point sticking therapy and TCM psychotherapy, and medication with deanxit all relieve depression after breast cancer operation. The efficacy of the combined program with auricular point sticking therapy and TCM psychotherapy involved is the best, and the efficacy of the auricular point sticking therapy is better than the oral administration of deanxit.</p>

[Effect of P38MAPK signal transduction pathway on apoptosis of THP-1 induced by allicin].

OBJECTIVE: The objective of this paper was to study the change of P38MAPK and Fas in the apoptosis of THP-1 cells induced by allicin. METHOD: The proliferation inhibition rates of THP-1 cells after various treatments were examined by MTT assay. Apoptosis rate was determined with Annexin V- FITC/PI double staining by flow cytometry. The expression and distribution change of the phosphorylation p38MAPK (P-p38MAPK) were detected by immunohistochemical staining. The changes of P-p38 MAPK and Fas proteins were detected by Western blot. RESULT: The proliferations of leukemia cell line THP-1 are inhibited by allicin. MTT assay showed that allicin can inhibit the proliferation of the THP-1 cell, and the inhibition was dependent on both dose and time. The IC50 of 72 hours was 12.8 mg x L(-1). Apoptosis rate detected by Annexin V-FITC/PI was proportional to the concentration of the allicin. After the immunohistochemical staining test, the P-p38MAPK was located in the cell nucleus and plasma, showing deep brown, when adding allicin to THP-1 cell. Western blot test showed that the P-p38MAPK proteins expression was proportional to the concentration of Allicin and was also dose dependent. The levels of P-p38MAPK in negative control group, 1/2 IC50 of 72 hours group and IC50 of 72 hours group were 0.259 8 +/- 0.013 2, 0.61 2 +/- 0.008 3 and 0.505 6 +/- 0.005 5 respectively, and the levels of Fas proteins were 0.287 4 +/- 0.008 9, 0.426 8 +/- 0.007 9 and 0.597 1 +/- 0.010 9 respectively. The difference was statistically significant when compared with the negative control group (P < 0.01). CONCLUSION: Allicin can significantly induce THP-1 cells apoptosis, and its mechanism may be related to the activation of P38MAPK/Fas.

[Apoptosis of U937 cell line promoted by matrine through MAPK signal transduction pathway].

OBJECTIVE: To study the anti-cancer effect of matrine (Mat) on U937 cell line and its possible molecular mechanism. METHOD: The cells were cultured in medium containing either 0.1, 0.2, 0.3, 0.4, or 0.5 g x L(-1) of Mat. The morphological alteration was observed by inverted microscopy and electron microscopy. Cell proliferation was analyzed by Try pan blue staining and MTT. The method of Western Blot was used to detect phosphorylation activity of MAPK. RESULT: Matrine had a significant inhibitory effect on proliferation of U937 cell line at the concentration of 0.2 g x L(-1). Treated with matrine of 0.2 g x L(-1) for 48 h, U937 cells became smaller and appeared more round than previously. The number of U937 cells showing apoptosis increased with elevation of the concentration of the matrine. Matrine had an ability of inhibiting the activity of ERK and increasing the activities of p38 and JNK to some degree in U937 cells. CONCLUSION: Matrine can inhibit the proliferation of U937 cell line in vitro and induce its apoptosis possibly through inhibiting the activity of ERK and increasing the activities of p38 and JNK in U937 cells.