The complete chloroplast genome sequence of Citrus maxima (Burm.) Merr. 'Guanximiyou'.

Citrus maxima (Burm.) Merr. 'Guanximiyou' is a major citrus tree largely cultivated in China. A previous study has reported the complete chloroplast genome of C. maxima, but there may be some differences between wild species and cultivating variety. In this study, the complete chloroplast genome sequence of 'Guanximiyou' pummelo was characterized using BGISEQ-500 sequencing. The chloroplast genome was 160,186 bp in length and separated into four distinct regions such as large single-copy region (87,939 bp), small single-copy region (18,395 bp), and a pair of inverted repeat regions (26,926 bp). The genome contained a total of 109 genes including 79 protein-coding genes, 29 tRNA genes, and 4 rRNA genes. Phylogenetic maximum-likelihood analysis revealed that 'Guanximiyou' pummelo was clustered with other Rutaceae species with high bootstrap values.

3D-QSAR, molecular docking, and molecular dynamics simulation of a novel thieno[3,4-d]pyrimidine inhibitor targeting human immunodeficiency virus type 1 reverse transcriptase.

Human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) is one of the most attractive drug targets for the treatment of AIDS. In this study, 67 thieno[3,4-d]pyrimidine derivatives were selected as novel HIV-1 RT inhibitors to combat viral resistance, and were subjected to 3 D-QSAR studies using CoMFA, CoMSIA, and T-CoMFA. In the 3 D-QSAR study, two methods of ligand-based alignment and pharmacophore-based alignment were used. The results showed that CoMFA (n = 8; q2 = 0.594; r2 = 0.974) and CoMSIA (n = 7; q2 = 0.528; r2 = 0.965) have good stability and predictability. The molecular docking study showed that the hydrogen bonding and van der Waals interactions of key residues such as Leu100, Lys101, Val106, Phe227 and Pro236 play an important role in ligand-receptor binding. Based on these results, 12 new thieno[3,4-d]pyrimidines were designed and their activities were predicted; the results indicated that these compounds have good predictive activity and reasonably good ADME/T profiles. MD simulation analysis of 50 ns showed that compound 23j formed four hydrogen bonds with the residues (Lys101, Lys104, Val106 and Thr318), and binds more closely to HIV-1 RT than compound 23j. Furthermore, the group at the R1 position and the horseshoe-like conformation of these compounds are critical for the inhibitory activity and stability. These results provide useful insights for the discovery and design of a new generation of HIV-1 RT inhibitors.Communicated by Ramaswamy H. Sarma.

Mycobacterium tuberculosis Infection Induces Low-Density Granulocyte Generation by Promoting Neutrophil Extracellular Trap Formation via ROS Pathway.

The roles and characteristics of low-density granulocytes (LDGs) have recently attracted attention; however, the mechanism of the formation of LDGs is yet unclear. In one of our previous studies, the frequency of LDGs was significantly elevated in the peripheral blood of tuberculosis patients, and in situ activation contributed to the generation of LDGs upon Mycobacterium tuberculosis infection. However, the underlying molecular mechanisms are yet to be elucidated. In the present study, the release of neutrophil extracellular traps (NETs) and the levels of ROS were regulated before the normal-density granulocytes (NDGs) to be infected with M. tuberculosis, and the conversion of NDGs to LDGs was monitored subsequently as well. The results showed that tuberculosis-related LDGs spontaneously released high levels of NETs. Promoting the release of NETs led to increase in the conversion of NDGs to LDGs in M. tuberculosis infection, while inhibiting the release of NETs suppressed this conversion after the infection. The M. tuberculosis infection significantly increased the ROS levels in neutrophils and the conversion of NDGs to LDGs. Scavenging ROS or blocking the ROS generation of M. tuberculosis-infected NDGs significantly suppressed the release of NETs and blocked the generation of LDGs. Moreover, inhibiting the formation of NETs without affecting the levels of ROS significantly decreased the conversion of NDGs to LDGs after M. tuberculosis infection. Overall, this study demonstrated that M. tuberculosis could induce the generation of LDGs by promoting the release of NET via ROS pathway.

Ursolic Acid Prevents Retinoic Acid-Induced Bone Loss in Rats.

OBJECTIVE: To examine the effects of ursolic acid (UA) on mitigating retinoic acid (RA)-induced osteoporosis in rats. METHODS: Fifty female Sprague-Dawley rats were randomly divided into the control group (n=10) and the osteoporosis group (n=40). The 40 osteoporosis rats were induced by 75 mg/(kg•d) RA once daily for 2 weeks, and then were randomly assigned to vehicle control (model), low-, middle-, and high-dose UA [(UA-L, UA-M, UA-H; 30, 60, 120 mg/(kg•d), respectively] groups (10 rats each). UA were administered once daily to the rats from the 3rd weeks for up to 4 weeks by gavage. Bone turnover markers [serum alkaline phosphatase (ALP), osteocalcin (OCN), urine deoxypyridinoline (DPD)] and other parameters, including serum calcium (S-Ca), serum phosphorus (S-P), urine calcium (U-Ca), urine phosphorus (U-P), and bone mineral density (BMD) of the femur, 4th lumbar vertebra and tibia, bone biomechanical properties and trabecular microarchitecture, were measured. RESULTS: The osteoporosis in rats was successfully induced by RA. Compared with the model group, UA-M and UA-H significantly reversed the RA-induced changes in S-P, U-Ca, U-P, ALP, OCN and urine DPD ratio and markedly enhanced the BMD of right femur, 4th lumbar vertebra and tibia (Plt;0.05 or Plt;0.01). Further, biomechanical test and microcomputed tomography evaluation also showed that UA-H drastically improved biomechanical properties and trabecular microarchitecture (Plt;0.05 or Plt;0.01). CONCLUSION: UA could promote bone formation, increase osteoblastic activity and reduce osteoclastic activity in rats, indicating that UA might be a potential therapeutic of RA-induced acute osteoporosis.

Gentiopicroside protects neurons from astrocyte-mediated inflammatory injuries by inhibition of nuclear factor-κB and mitogen-activated protein kinase signaling pathways.

The effects of gentiopicroside (Gent), an active component derived from the traditional Chinese medicine Gentiana macrophylla, on lipopolysaccharide-induced astrocyte activation and subsequent neuronal damage were investigated. Gent significantly inhibited the release of tumor necrosis factor-α, interleukin-1ß, nitric oxide, and prostaglandin E, as well as expressions of inducible nitric oxide synthase and cyclooxygenase-2 in lipopolysaccharide-induced primary astrocytes. Furthermore, Gent relieved neurotoxicity from astrocyte-mediated inflammatory injury. Mechanism studies indicated that Gent significantly suppressed nuclear factor-κB nuclear translocation and down-regulated c-Jun-N-terminal kinase/stress-activated protein kinase mitogen-activated protein kinase phosphorylation levels with little influence on elevated p-p38 levels. Taken together, our findings suggested Gent could prevent the neurotoxicity related to astrocyte-mediated inflammatory injury by inhibition of nuclear factor-κB and mitogen-activated protein kinase signaling pathways. The study also indicated that neuronal injury could be prevented by promptly modulating inflammatory responses of astrocytes.

Gentiopicroside abrogates lipopolysaccharide-induced depressive-like behavior in mice through tryptophan-degrading pathway.

Targeting neuroinflammatory disturbances has been acknowledged as a potential strategy for treatment of depressive disorder in humans. Over-activation of tryptophan-degrading pathway by pro-inflammatory cytokines resulted in N-methyl-d-aspartate (NMDA)-mediated excitotoxicity, which is implicated in pathophysiology of depression. Gentiopicroside (Gent) has powerful anti-inflammatory property and exhibits promising antidepressant effect in an animal model of pain/depression dyad by down-regulating GluN2B-containing NMDA receptors. Therefore, the present study aimed to investigate the ability of Gent to abolish depressive-like behavior induced by lipopolysaccharide (LPS) in mice. Acute administration of LPS (0.5 mg/kg, i.p.) increased immobility time in both forced swimming test (FST) and tail suspension test (TST) without affecting spontaneous locomotor activity, indicative of depressive-like behavior. Gent (50 mg/kg, i.p.) administered once a day for three consecutive days prevented the development of depressive-like behavior induced by LPS. The antidepressant-like effect was paralleled with restoration of LPS-induced alterations in brain inflammatory mediators (i.e. IL-1ß and TNF-α). In addition, Gent prevented over-activation of indoleamine 2,3-double oxygen enzyme (IDO) and recovered GluN2B subunit expression in the PFC challenged by LPS. In conclusion, our results suggested that Gent pretreatment provided protection against LPS-induced depressive-like behavior and the effect appeared to be demonstrated, at least partially, by blocking various steps of tryptophan-degrading pathway.

Echinocystic acid reduces reserpine-induced pain/depression dyad in mice.

Chronic pain has consistently been correlated with depression. Echinocystic acid (EA), a natural triterpone enriched in various herbs and used for medicinal purpose in many Asian countries, exhibits anti-inflammatory and analgesic activities. However, little is known the effects of EA on the depression. In present study, we investigated the anti-depression activities in the mouse model of reserpine-induced pain-depression dyad. Reserpine (1 mg/kg subcutaneously daily for 3 days) caused significant depression-like behaviors and pain sensation. Subsequent treatment of EA (5 mg/kg intragastrically daily for 5 days) attenuated the reserpine-induced pain/depression dyad as shown by the increase of pain threshold and the behaviors in forced swimming test, tail suspension test, and open field test. Furthermore, treatment of EA reversed the decrease of biogenic amines (norepinephrine, dopamine, and serotonin) in the brain region of hippocampus, a structure involved in the formation of emotional disorders. Levels of serotonin receptor 5-HT1A were decreased and levels of 5-HT2A were increased in the reserpine-injected mice. Treatment of EA could restore the alterations of serotonin receptors. At the same time, the increase in GluN2B-containing NMDA receptors, p-GluA1-Ser831, PSD-95 and CaMKII were integrated with the increase in caspase-3 and iNOS levels in the hippocampus of the reserpine-injected mice. EA significantly reversed the changes of above proteins. However, EA did not affect the levels of GluN2A-containing NMDA receptors and the total levels of GluA1 and p-GluA1-Ser845. Our study provides strong evidence that EA attenuates reserpine-induced pain/depression dyad partially through regulating the biogenic amines levels and GluN2B receptors in the hippocampus.

Osteoprotective Effect of Echinocystic Acid, a Triterpone Component from Eclipta prostrata, in Ovariectomy-Induced Osteoporotic Rats.

Echinocystic acid (EA) is a natural triterpone enriched in various herbs and has been used for medicinal purposes in China. In the present study, we systematically examined the effects of EA on ovariectomy-induced osteoporosis in rats for the first time. Three-month-old female ovariectomy (OVX) Sprague-Dawley rats were used to evaluate the osteoprotective effect of EA. Results showed that administration of EA (5 or 15 mg/kg/day) for 12 weeks prevented lower levels of maximum stress and Young's modulus of femur induced by OVX. EA also recovered bone metabolic biomarkers levels in OVX rats, including osteocalcin, alkaline phosphatese, deoxypyridinoline, and urinary calcium and phosphorus. EA (5 and 15 mg/kg/day) could prevent the alteration of total bone mineral density in the femur caused by OVX. However, only high dose (15 mg/kg/day) of EA significantly improved trabecular architecture, as evidenced by higher levels of bone volume/tissue volume, trabecula number, and trabecula thickness, and lower levels of trabecula separation and structure model index compared with OVX rats. In addition, EA treatment decresed the serum levels of IL-1ß and TNF-α in OVX rats. In conclusion, EA could prevent reduction of bone mass and strength and improve the cancellous bone structure and biochemical properties in OVX rats. Hence, EA may serve as a new candidate or a leading compound for anti-osteoporosis.

Osteoprotective effects of estrogen membrane receptor GPR30 in ovariectomized rats.

G protein-coupled estrogen receptor 30 (GPR30) is expressed in bone tissue. However, little is known regarding the function of GPR30 in postmenopausal osteoporosis. In this study, we examined the effects of GPR30 on ovariectomy (OVX)-induced osteoporosis in rats, including the effects on proliferation, differentiation, and expression of proteins in osteoblasts. Administration of G1 (35 µg/kg, ip, 3 times/week for 6 weeks), a specific agonist of GPR30, prevented OVX-induced increase in bone turnover rate, decrease in bone mineral content and bone mineral density, damage to bone structure, and aggravation of bone biomechanical properties. In addition, G1 did not affect uterine weight in the OVX rats. Osteoblasts isolated from calvarias from newborn rats were used to explore the underlying mechanisms. G1 (150 pM) promoted proliferation and differentiation of osteoblasts through a positive feedback of GPR30, which then activated the PI3K-Akt, ERK, and CREB pathways. G15 (750 pM), a specific antagonist of GPR30, reversed the above effects initiated by G1 treatment. In conclusion, activation of GPR30 protected bones against osteoporosis in OVX rats and exerted no untoward effect on the uterus. We suggest that GPR30 can be used as an effective therapeutic target for the prevention and treatment of postmenopausal osteoporosis.

[Mass spectrometric analysis of quinazoline alkaloid and limonin from Fructus Evodiae].

OBJECTIVE: To study Mass spectrometry of quinazoline alkaloid and limonin from Fructus Evodiae by an electrospray ionization (ESI) tandem mass spectrometry. METHODS: The ESI source was set at the positive ionization mode, and the MS1 and MS2 spectra of quinazoline alkaloid and limonin were acquired. RESULTS: The main fragments of quinazoline alkaloid and limonin from Fructus Evodiae were analyzed. CONCLUSIONS: The profiles of fragments will be usefull to set quality criteria and to study chemical composition of Fructus Evodiae.

[Analysis of the main components of coptis-evodia herb couple by HPLC-DAD-MS].

A comprehensive HPLC-DAD-MS method was developed to study the chemical components of semi-bionic extract of Coptis-Evodia herb couple. The extract was isolated on a Hypersil BDS C18 column (4.6 mm x 200 mm, 5 microm) using acetonitrile-ammonium formic buffer as mobile phase by gradient elution. Detection was performed on DAD and MS equipped with an electrospray ionization (ESI) source by full scan and product full scan on positive mode. The chromatogram of Coptis-Evodia showed seventeen main peaks, eight of which were from Evodia while the others were from Coptis. By comparison of the retention time, the on-line UV spectra and MS spectra, four peaks were identified as jatrorrhizine, hydroxevodiamine, palmatine and berberine, and three peaks were deduced as epiberberine, columbamine and coptisine. In addition, berberine and palmatine were quantitatively determined. No new component was created in the semi-bionic extract of the herb couple, yet the solubilities of berberine and palmatine decreased.