Minor Hydroxylated Triterpenoids Produced in Engineered Yeast by the Enzymes OSC and CYP716s from the Plant Enkianthus chinensis and Their Anti-Inflammatory and Hepatoprotective Activities.

Triterpenoids are a type of specialized metabolites that exhibit a wide range of biological activities. However, the availability of some minor triterpenoids in nature is limited, which has hindered our understanding of their pharmacological potential. To overcome this limitation, heterologous biosynthesis of triterpenoids in yeast has emerged as a promising and time-efficient production platform for obtaining these minor compounds. In this study, we analyzed the transcriptomic data of Enkianthus chinensis to identify one oxidosqualene cyclase (EcOSC) gene and four CYP716s. Through heterologous expression of these genes in yeast, nine natural pentacyclic triterpenoids, including three skeleton products (1-3) produced by one multifunctional OSC and six minor oxidation products (4-9) catalyzed by CYP716s, were obtained. Of note, we discovered that CYP716E60 could oxidize ursane-type and oleanane-type triterpenoids to produce 6ß-OH derivatives, marking the first confirmed C-6ß hydroxylation in an ursuane-type triterpenoid. Compound 9 showed moderate inhibitory activity against NO production and dose-dependently reduced IL-1ß and IL-6 production at the transcriptional and protein levels. Compounds 1, 2, 8, and 9 exhibited moderate hepatoprotective activity with the survival rates of HepG2 cells from 61% to 68% at 10 µM.

Prenylated C6-C3 derivatives from the root of Illicium ternstroemioides with antiviral activity.

Six previously undescribed prenylated C6-C3 derivatives (1-6) were isolated from the root of Illicium ternstroemioides A. C. Smith. Their structures were elucidated based on extensive spectroscopic analyses (UV, IR, 1D and 2D NMR, and HRESIMS). The absolute configurations of 1-3 were determined using electronic circular dichroism (ECD), and Mo2(OAc)4 induced circular dichroism (ICD). Compound 3 exhibited weak activity against Coxsackievirus B3 with an IC50 value of 33.3 µM, and compound 5 exhibited more potent activity against Coxsackievirus B3 with an IC50 value of 6.4 µM.

Cadinane sesquiterpenes from the stems and branches of Illicium ternstroemioides.

Three new cadinane sesquiterpenes (1-3) and three known sesquiterpenes were isolated from the stems and branches of Illicium ternstroemioides A. C. Smith. The structures of the new compounds were elucidated by extensive analysis of spectroscopic and HRESIMS data. The structures of illiternins A-C (1-3) were confirmed by single crystal X-ray diffraction, allowing for the determination of their absolute configurations. Compounds 3 and 6 exhibited antiviral activity against Coxsackievirus B3 with IC50 values of 33.3 and 57.7 µM, respectively.

Characterization and Engineering of Two Highly Paralogous Sesquiterpene Synthases Reveal a Regioselective Reprotonation Switch.

Sesquiterpene synthases (STPSs) catalyze carbocation-driven cyclization reactions that can generate structurally diverse hydrocarbons. The deprotonation-reprotonation process is widely used in STPSs to promote structural diversity, largely attributable to the distinct regio/stereoselective reprotonations. However, the molecular basis for reprotonation regioselectivity remains largely understudied. Herein, we analyzed two highly paralogous STPSs, Artabotrys hexapetalus (-)-cyperene synthase (AhCS) and ishwarane synthase (AhIS), which catalyze reactions that are distinct from the regioselective protonation of germacrene A (GA), resulting in distinct skeletons of 5/5/6 tricyclic (-)-cyperene and 6/6/5/3 tetracyclic ishwarane, respectively. Isotopic labeling experiments demonstrated that these protonations occur at C3 and C6 of GA in AhCS and AhIS, respectively. The cryo-electron microscopy-derived AhCS complex structure provided the structural basis for identifying different key active site residues that may govern their functional disparity. The structure-guided mutagenesis of these residues resulted in successful functional interconversion between AhCS and AhIS, thus targeting the three active site residues [L311-S419-C458]/[M311-V419-A458] that may act as a C3/C6 reprotonation switch for GA. These findings facilitate the rational design or directed evolution of STPSs with structurally diverse skeletons.

seco-Sesquiterpenes and acorane-type sesquiterpenes with antiviral activity from the twigs and leaves of Illicium henryi Diels.

Chemical investigation of an alcohol extract from the twigs and leaves of Illicium henryi Diels resulted in the isolation of two new acorane-related seco-sesquiterpenes (1 and 3), two new acorane-related seco-norsesquiterpenes (2 and 4), one new 2-epi-cedrane sesquiterpene (5), eight new acorane-type sesquiterpenes (6-13), and a known major constituent of acorenone B (14). Their structures were established by interpreting extensive spectroscopic data, including HRESIMS, NMR (1H and 13C NMR, 1H-1H COSY, HSQC, and HMBC), and NOE difference spectra analysis. The absolute configurations of 1, 2, 4-7, 9, 10, and 14 were determined by X-ray crystallography, while chemical transformation methods were performed with compound 14 as the starting material to elegantly solve the absolute configuration issue of compounds 8 and 11-13. Notably, 1 and 2 are seco-sesquiterpenes that are related to acorane and possess an unusual ketal-linked hemiacetal in a 6,8-dioxabicyclo[3.2.1]octan-7-ol scaffold ring system. Plausible biosynthetic pathways for compounds 1-14, which were derived from the acorane skeleton, were proposed. All the isolated compounds (1-14) were evaluated for their antiviral and cytotoxic activities.

Minor terpenoids from the leaves of Craibiodendron yunnanense.

One new taraxastane-type triterpenoid, three new grayanane-type diterpenoids (2 - 4), and 12 known compounds (5 - 16) were isolated from the leaves of Craiobiodendron yunnanens W. W. Smith. The structures of these compounds were elucidated on the basis of their spectroscopic data and chemical evidence. Compounds 1 and 8 exhibited partly anti-inflammatory activity based on the inhibition of NF-κB activity in SW480 cells at 10 µM with inhibition ratios of 60.53 and 59.20%, respectively. Compounds 10 and 13 showed excellent cytotoxicity against human leukemia cell (MV4-11) at 10 µM with inhibition ratios of 43.02 and 49.11%, respectively.

[Chemical constituents from leaves of Craibiodendron yunnanense].

The present study investigated the chemical constituents from the leaves of Craibiodendron yunnanense. The compounds were isolated and purified from the leaves of C. yunnanense by a combination of various chromatographic techniques including column chromatography over polyamide, silica gel, Sephadex LH-20, and reversed-phase HPLC. Their structures were identified by extensive spectroscopic analyses including MS and NMR data. As a result, 10 compounds, including melionoside F(1), meliosmaionol D(2), naringenin(3), quercetin-3-O-α-L-arabinopyranoside(4), epicatechin(5), quercetin-3'-glucoside(6), corbulain Ib(7), loliolide(8), asiatic acid(9), and ursolic acid(10), were isolated. Compounds 1 and 2 were two new compounds, and compound 7 was isolated from this genus for the first time. All compounds showed no significant cytotoxic activity by MTT assay.

CT Scans and Cancer Risks: A Systematic Review and Dose-response Meta-analysis.

BACKGROUND: There is still uncertainty on whether ionizing radiation from CT scans can increase the risks of cancer. This study aimed to identify the association of cumulative ionizing radiation from CT scans with pertaining cancer risks in adults. METHODS: Five databases were searched from their inception to November 15, 2020. Observational studies reporting cancer risks from CT scans in adults were included. The main outcome included quantified cancer risks as cancer case numbers in exposed/unexposed adult participants with unified converted measures to odds ratio (OR) for relative risk, hazard ratio. Global background radiation (2.4 mSv per year) was used as control for lifetime attribution risk (LAR), with the same period from incubation after exposure until survival to 100 years. RESULTS: 25 studies were included with a sum of 111,649,943 participants (mean age: 45.37 years, 83.4% women), comprising 2,049,943 actual participants from 6 studies with an average follow-up period as 30.1 years (range, 5 to 80 years); 109,600,000 participants from 19 studies using LAR. The cancer risks for adults following CT scans were inordinately increased (LAR adults, OR, 10.00 [95% CI, 5.87 to 17.05]; actual adults, OR, 1.17 [95%CI, 0.89 to 1.55]; combined, OR, 5.89 [95%CI, 3.46 to 10.35]). Moreover, cancer risks elevated with increase of radiation dose (OR, 33.31 [95% CI, 21.33 to 52.02]), and multiple CT scan sites (OR, 14.08 [95% CI, 6.60 to 30.05]). The risk of solid malignancy was higher than leukemia. Notably, there were no significant differences for age, gender, country, continent, study quality and studying time phrases. CONCLUSIONS: Based on 111.6 million adult participants from 3 continents (Asia, Europe and America), this meta-analysis identifies an inordinately increase in cancer risks from CT scans for adults. Moreover, the cancer risks were positively correlated with radiation dose and CT sites. The meta-analysis highlights the awareness of potential cancer risks of CT scans as well as more reasonable methodology to quantify cancer risks in terms of life expectancy as 100 years for LAR. PROSPERO TRIAL REGISTRATION NUMBER: CRD42019133487.

Six new sesquiterpenoids from the fruits of Xanthium italicum.

Four new germacrane-type sesquiterpenoids (1-4) and two new guaiane-type sesquiterpenoids (5-6) were isolated from the fruits of Xanthium italicum Moretti. The structures of the new compounds were elucidated on the basis of spectroscopic analysis and X-ray diffraction experiment. Compounds 1, 2 and 6 showed the anti-inflammatory effects against the activation of NF-κB induced by lipopolysaccharide (LPS) with IC50 values of 20.12, 22.89 and 68.66 µM, respectively.

New lignans and diterpenoid glycosides from the fruits of Xanthium italicum Moretti.

A pair of new lignans [(+)- 1 and (-)- 1] and three new compounds (2-4), together with a known compound 5, were isolated from the fruits of Xanthium italicum Moretti. The structures of these compounds were determined on the basis of spectroscopic analysis, particularly HR-ESI-MS and 1 D and 2 D NMR. Compounds 2 and 3 showed antinociceptive effects in an acetic acid-induced writhing test in mice with the writhe inhibition rates of 80.50% and 67.89% at the dose of 20 mg/kg, respectively.

Minor terpenoids from the stems and twigs of Rhododendron ovatum.

Four minor undescribed terpenoids, including a monoterpenoid (1) and three triterpenoids (3, 6 and 7), together with 26 known terpenoids were isolated from the stems and twigs of Rhododendron Ovatum. Their structures were identified by extensive spectroscopic analyses and electronic circular dichroism (ECD) techniques. Compound 10 showed excellent cytotoxicity against human colon cancer cell (HCT-116) with IC50 value of 2.56 µM. Compounds 9 and 19 exhibited partly inhibitory effects on nitric oxide production stimulated by lipopolysaccharide-induced neuroinflammation in microglia cells at 10 µM with inhibition ratios of 39.70% and 28.08%, respectively.

Oxygenated pentacyclic triterpenoids from the stems and branches of Enkianthus chinensis.

Thirty new pentacyclic triterpenoids, including five oleanane-type (1-5), twenty-three ursane-type (9-23, 26-33) and two taraxerane-type (24 and 25), along with fourteen known triterpenoids, were isolated from the stems and branches of Enkianthus chinensis. Their structures were elucidated by extensive spectroscopic analyses, X-ray crystallographic data and electronic circular dichroism (ECD) techniques. Sixteen compounds (1-5, 9-13, 20, 22, 32, 34-36) bearing a gem-hydroxymethyl group at C-4 represent rare examples of pentacyclic triterpenoids. In the in vitro biological activity evaluation, compounds 8, 9, 12-14, 17, 24, and 44 exhibited potent hepatoprotective effects at 10 µM. Moreover, compound 25 showed latent activity against HSV-1 with an IC50 value of 6.4 µM.

Low Radiation X-rays: Benefiting People Globally by Reducing Cancer Risks.

Modern medical imaging facilitates the diagnosis and treatment of human diseases. However, few people are aware of the cons of radiation exposure from medical imaging. Emerging evidence reveals that cumulative doses of radiation exposure will increase the morbidity and mortality of pertaining cancer. As a special young population, patients with adolescent idiopathic scoliosis (AIS) suffer more radiation harms from repeated diagnostic imaging, most of which can be avoided in clinical practice. Accumulating evidence highlights reduced cancer risks of radiation exposure for AIS patients with low/zero radiation imaging modalities proposed, amongst which easy conversion from anterior-posterior (AP) to posterior-anterior (PA) projection for whole-spine radiographs should be stressed. It can greatly reduce radiation doses without compromising the quality of diagnostic imaging. Tight collimation combined with PA projection can further reduce radiation harms, and need to be spread to benefit people globally.

Hydroxylated Ethacrylic and Tiglic Acid Derivatives from the Stems and Branches of Enkianthus chinensis and Their Potential Anti-inflammatory Activities.

Two new hydroxylated ethacrylic acid derivatives (compounds 1 and 2) and 11 new hydroxylated tiglic acid derivatives (compounds 3-13), together with one known compound (compound 14), were isolated from the stems and branches of Enkianthus chinensis. Their structures were established by extensive spectroscopic analyses, while their absolute configurations were determined by X-ray crystallographic methods (compounds 1 and 2), Mo2(OAc)4-induced electronic circular dichroism experiments (compounds 3 and 4), and chemical methods (compounds 5-11). This study is the first investigation on the secondary metabolites of this species. The anti-inflammatory activities of all isolated compounds were evaluated in an LPS-induced mouse peritoneal macrophage model. Notably, compounds 3 and 12 both exerted potent inhibitory effects on NO production with IC50 values of 2.9 and 1.2 µM, respectively.

Cancer and mortality risks of patients with scoliosis from radiation exposure: a systematic review and meta-analysis.

PURPOSE: The study aimed for unraveling the long-term health impact of cumulative radiation exposure from full-spine radiographs on children/adolescents with scoliosis. METHODS: All cohort, case-control or cross-sectional studies about radiation exposure to scoliosis patients with follow-up period as 20 years or more were included. Meta-analyses were performed for outcomes reported in two or more studies. RESULTS: A total of 9 eligible studies involving 35,641 participants between 1912 and 1990 fulfilled the inclusion criteria, including 18,873 patients with scoliosis and 16,768 controls as regional matched general population. The average number of full-spine radiographs was 23.13 (range: 0-618) according to 14,512 patients between 1912 and 1990 in five studies. The estimated mean cumulated radiation dose of breast was 11.35 cGy. In comparison with controls, pooled incidence rates of cancer, breast cancer and cancer mortality of patients with scoliosis were statistically significant higher [rate of cancer, odds risk (OR) = 1.46, p < 0.00001; breast cancer, OR = 1.20, p = 0.02; cancer mortality, OR = 1.50, p < 0.00001]. No statistically significant differences were found in terms of reproductive events for scoliosis patients, pulmonary function and physical activity for adolescent idiopathic scoliosis patients. CONCLUSIONS: Based on 35,641 participants with over 20 years' observations from 1912 to 1990, repeated radiographs and pertaining cumulative radiation dose resulted in elevated rates of cancer, breast cancer and cancer mortality for children/adolescents with scoliosis in comparison with matched general population. It is recommended that low-radiation or radiation-free and efficient methods should be used to monitor the evolution of children/adolescents with scoliosis.

[Analysis of chemical constituents in Shenmai Injection by LC-Q-TOF-MS and LC-IT-MS].

The chemical constituents in Shenmai Injection(SMI) were qualitatively analyzed by using liquid chromatography/quadrupole time-of-flight mass spectrometry(LC-Q-TOF-MS) and liquid chromatography-ion trap-mass spectrometry(LC-IT-MS). The analysis was performed on an Agilent Zorbax SB-C_(18)(4.6 mm×250 mm, 5 µm) and gradient elution was carried out with 0.05% formic acid solution-acetonitrile as mobile phase at a flow rate of 0.6 mL·min~(-1) and a column temperature of 30 ℃. Mass spectrometry data of the components in SMI were collected in negative ion mode. The structures of components were speculated and identified by analyzing mass spectrometry data, comparing with standards, and referring to related literature. A total of 64 components in SMI were estimated, and the structures were confirmed in 16 of them by comparison with standards. Fifty-six compounds derived from Ginseng Radix et Rhizoma Rubra included 34 protopanaxadiol ginsenosides, 19 protopanaxatriol ginsenosides, 1 oleanane ginsenosides and 2 other glycosides. Eight compounds derived from Ophiopogonis Radix included 7 steroidal saponins, and 1 monoterpene glycoside. The results of this study would provide an important theoretical basis for the improvement of the quality control standards and the discovery of effective constituents in SMI.

Cytotoxic xanthanolide sesquiterpenes from the fruits of Xanthium italicum Moretti.

One previously undescribed xanthanolide sesquiterpene dimer pungiolide P (1), possessing an unprecedented scaffold with a 5/7/5/7/5 ring system skeleton and its intermediate pungiolide Q (2), ten xanthanolide sesquiterpenes (3-12), two eudesmene sesquiterpene derivatives (13-14), one phenylpropionic acid derivative (15), together with eleven known compounds (16-26) were obtained from the fruits of Xanthium italicum Moretti. A possible biosynthetic pathway for pungiolide P (1) was also proposed, which was supported by its bio-synthetic intermediate (2). Compounds 1, 4-5, 18-21, and 25 exhibited cytotoxic activity against a variety of human cancer cell lines. Furthermore, compounds 1, 4-5, could cause blockage of the cell cycle in the G2/M phase and induce apoptosis in H460 cells. Notably, pungiolide P (1) exhibited significantly superior cytotoxicity compared to previously reported compounds, providing valuable insights for natural anti-tumor sources.

CK8 phosphorylation induced by compressive loads underlies the downregulation of CK8 in human disc degeneration by activating protein kinase C.

Cytokeratin 8 (CK8) is a member of the cytokeratins family with multiple functions on the basis of its unique structural hallmark. The aberrant expression of CK8 and its phosphorylation are pertinent with various diseases. We have previously shown that CK8 exists in normal human nucleus pulposus (NP) cells and decreases as the intervertebral disc degenerates. However, the underlying molecular regulatory machinery of CK8 in intervertebral disc degeneration (IDD) has not been clarified. Here, we collected NP samples from patients with idiopathic scoliosis as control and IDD as degenerate groups. We found that CK8 expression decreased in IDD with an increased phosphorylation in degenerate NP cells. Moreover, NP cells were cultured under different compressive load schemes for diverse time duration. We found that compressive loads resulted in phosphorylation and disassembly of CK8 in a time-dependent and degree-dependent manner in vitro. The activation of protein kinase C was a significant molecular factor contributing to this phenomenon. Taken together, this study is the first to address the molecular mechanisms of CK8 downregulation in NP cells. Importantly, our findings provide clues regarding a molecular link between compressive loads and CK8 alterations, which shed a novel light on the etiology of IDD.