Alpha-Asarone Ameliorates Neurological Dysfunction of Subarachnoid Hemorrhagic Rats in Both Acute and Recovery Phases via Regulating the CaMKII-Dependent Pathways.

Early brain injury (EBI) is the leading cause of poor prognosis for patients suffering from subarachnoid hemorrhage (SAH), particularly learning and memory deficits in the repair phase. A recent report has involved calcium/calmodulin-dependent protein kinase II (CaMKII) in the pathophysiological process underlying SAH-induced EBI. Alpha-asarone (ASA), a major compound isolated from the Chinese medicinal herb Acorus tatarinowii Schott, was proven to reduce secondary brain injury by decreasing CaMKII over-phosphorylation in rats' model of intracerebral hemorrhage in our previous report. However, the effect of ASA on SAH remains unclear, and the role of CaMKII in both acute and recovery stages of SAH needs further investigation. In this work, we first established a classic SAH rat model by endovascular perforation and intraperitoneally administrated different ASA doses (10, 20, and 40 mg/kg) 2 h after successful modeling. Then, the short- and long-term neurobehavioral performances were blindly evaluated to confirm ASA's efficacy against SAH. Subsequently, we explored ASA's therapeutic mechanism in both acute and recovery stages using histopathological examination, TUNEL staining, flow cytometry, Western-blot, double-immunofluorescence staining, and transmission electron microscopy (TEM) observation. Finally, KN93, a selective CaMKII inhibitor, was applied in oxyhemoglobin-damaged HT22 cells to explore the role of CaMKII in ASA's neuroprotective effect. The results demonstrated that ASA alleviated short- and long-term neurological dysfunction, reduced mortality and seizure rate within 24 h, and prolonged 14-day survival in SAH rats. Histopathological examination showed a reduction of neuronal damage and a restoration of the hippocampal structure after ASA treatment in both acute and recovery phases of SAH. In the acute stage, the Western-blot and flow cytometer analyses showed that ASA restored E/I balance, reduced calcium overload and CaMKII phosphorylation, and inhibited mitochondrion-involved apoptosis, thus preventing neuronal damage and apoptosis underlying EBI post-SAH. In the recovery stage, the TEM observation, double-immunofluorescence staining, and Western-blot analyses indicated that ASA increased the numbers of synapses and enhanced synaptic plasticity in the ipsilateral hippocampi, probably by promoting NR2B/CaMKII interaction and activating subsequent CREB/BDNF/TrkB signaling pathways. Furthermore, KN93 notably reversed ASA's neuroprotective effect on oxyhemoglobin-damaged HT22 cells, confirming CaMKII a potential target for ASA's efficacy against SAH. Our study confirmed for the first time that ASA ameliorated the SAH rats' neurobehavioral deterioration, possibly via modulating CaMKII-involved pathways. These findings provided a promising candidate for the clinical treatment of SAH and shed light on future drug discovery against SAH.

Effect of zero-valent iron nanoparticles on taxonomic composition and hydrogen production from kitchen waste.

This study investigated the effects of varying zero-valent iron (ZVI) (0 to 5,000 mg/L) on fermentative hydrogen (H2) production, metabolic pattern, and taxonomic profile by using kitchen waste as substrate. The study demonstrated that the supplementation of 500 mg ZVI/L resulted in the highest H2 yield (219.68 ± 11.19 mL H2/g-volatile solids (VS)added), which was 19% higher than the control. The metabolic pattern analysis showed that acetic and butyric acid production primarily drove the H2 production. The taxonomic analysis further revealed that Firmicutes (relative abundance (RA): 80-96%) and Clostridium sensu stricto 1 (RA: 68-88%) were the dominant phyla and genera, respectively, during the exponential gas production phase, supporting the observation of accumulation of acetic and butyric acids. These findings suggest that supplementation of ZVI can enhance H2 production from organic waste and significantly influence the metabolic pattern and taxonomic profile, including the metalloenzymes.

Clinical outcomes of diode laser as an adjunct to nonsurgical periodontal therapy for residual periodontal pockets in mandibular second molars-a randomized controlled clinical trial.

OBJECTIVES: The aim of this study was to evaluate the clinical outcomes of diode laser as an adjunct to nonsurgical periodontal therapy (NSPT) for residual periodontal pockets in mandibular second molars. MATERIALS AND METHODS: Sixty-seven mandibular second molars (154 residual periodontal pockets) were recruited into the study and randomly assigned to the Laser + NSPT group and the NSPT group. The Laser + NSPT group underwent NSPT adjunct with diode laser radiation (wavelength: 810 nm, power: 1.5 W, 40 s maximum), while the NSPT group underwent nonsurgical periodontal therapy alone. Clinical parameters were measured at baseline (T0) and 4(T1), 12(T2), and 24(T3), weeks after treatment. RESULTS: Periodontal pocket depth (PPD), clinical attachment loss (CAL), and bleeding on probing (BOP) in both groups showed significant improvements at the end of study compared to baseline. The reductions of PPD, CAL, and BOP in the Laser + NSPT group were significantly greater than NSPT group. At T3, the Laser + NSPT group had a mean PPD of 3.06 ± 0.86 mm, CAL of 2.58 ± 0.94 mm and BOP of 15.49%, while the NSPT group had a mean PPD of 4.46 ± 1.57 mm, CAL of 3.03 ± 1.25 mm and BOP of 64.29%. CONCLUSIONS: The diode laser as an adjunct to nonsurgical periodontal therapy may contribute to clinical outcomes for residual periodontal pockets. However, the approach may cause reduction of keratinized tissue width. TRIAL REGISTRATION NUMBER: This study was registered in the Chinese Clinical Trial Registry ChiCTR2200061194. CLINICAL RELEVANCE: Diode laser as an adjunct to nonsurgical periodontal therapy may contribute to the clinical outcomes for residual periodontal pockets in mandibular second molars.

Psychological stress: neuroimmune roles in periodontal disease.

Periodontitis is a chronic inflammatory disease that starts with pathogenic bacteria and is mediated by a combination of multiple factors. Psychosomatic factors are considered to be one of the most critical risk factors for periodontal disease. Psychological stress may threaten periodontal immune homeostasis in multiple ways by affecting the hypothalamic-pituitary-adrenal cortex system, the locus ceruleus-sympathetic-adrenal medulla system, and the peptidergic nervous system. In this review, we outline the complex role of psychological stress in promoting the development of periodontal disease, focusing on the effects of stress on flora metabolism, tissue inflammation, and alveolar bone homeostasis. At the same time, we broadly and deeply summarize the potential mechanisms of psychological stress-induced periodontal disease, emphasize the importance of neuroimmune modulation for periodontal health, and expect to provide a new perspective for periodontal science based on psychoneuroimmunology.

Emerging trends in the pretreatment of microalgal biomass and recovery of value-added products: A review.

Microalgae are a promising source of raw material (i.e., proteins, carbohydrates, lipids, pigments, and micronutrients) for various value-added products and act as a carbon sink for atmospheric CO2. The rigidity of the microalgal cell wall makes it difficult to extract different cellular components for its applications, including biofuel production, food and feed supplements, and pharmaceuticals. To improve the recovery of products from microalgae, pretreatment strategies such as biological, physical, chemical, and combined methods have been explored to improve whole-cell disruption and product recovery efficiency. However, the diversity and uniqueness of the microalgal cell wall make the pretreatment process more species-specific and limit its large-scale application. Therefore, advancing the currently available technologies is required from an economic, technological, and environmental perspective. Thus, this paper provides a state-of-art review of the current trends, challenges, and prospects of sustainable microalgal pretreatment technologies from a microalgae-based biorefinery concept.

Alpha-asarone ameliorates neurological deterioration of intracerebral hemorrhagic rats by alleviating secondary brain injury via anti-excitotoxicity pathways.

BACKGROUND: Secondary brain injury (SBI) has been confirmed as a leading cause for the poor prognosis of patients suffering from intracerebral hemorrhage (ICH). SBI co-exists in ischemia and hemorrhagic stroke. Neuro-excitotoxicity is considered the initiating factor of ICH-induced SBI. Our previous research has revealed alpha-asarone (ASA)'s efficacy against cerebral ischemia-reperfusion stroke by mitigating neuro-excitotoxicity. It is not yet known if ASA exhibit neuroprotection against ICH. PURPOSE: This work aimed to investigate ASA's therapeutic effects and potential mechanisms of action against ICH in a classic rat model induced by collagenase Ⅶ injection. METHODS: An in vivo ICH model of Sprague-Dawley rats was established by collagenase Ⅶ injection. We administrated different ASA doses (10, 20, or 40 mg/kg, i.p.) at 2 h post-ICH. Then, rats' short- and long-term neurobehavioral function, bodyweight change, and learning and memory ability were blindly evaluated. Histological, Nissl, and flow cytometry were applied to assess the neuronal damage post-ICH. The wet/dry method and Evans blue extravasation estimated brain edema and blood-brain barrier function. Pathway-related proteins were investigated by immunofluorescence staining, enzyme-linked immunosorbent assay, and Western-blot analysis. RESULTS: The results demonstrated that ASA ameliorated neurological deterioration, bodyweight loss, and learning and memory ability of ICH rats. Histological, Nissl, and flow cytometry analyses showed that ASA reduced neuronal damage and apoptosis post-ICH. Besides, ASA probably mitigated brain edema and blood-brain barrier dysfunction via inhibiting astrocyte activation and consequent pro-inflammatory response. The mechanism investigation attributed ASA's efficacy to the following aspects: 1) promoting sodium ion excretion, thus blocking excitatory signal transduction along the axon; 2) preventing glutamate-involved pathways, i.e., decrease of N-methyl-d-aspartic acid receptor subunit 2B, increase of glutamate transporter-1, and alleviation of calcium-related cascades, mitochondrion-associated apoptosis, and neuronal autophagy; 3) enhancing the expression of GABAARs, thus abating neuronal excitotoxicity. CONCLUSION: Our study first confirmed the effect of ASA on ameliorating the neurobehavioral deterioration of ICH rats, possibly via alleviation of glutamate-involved neuro-excitotoxicity, i.e., calcium cascades, mitochondrion-involved apoptosis, neuronal autophagy, and astrocyte-related inflammation. These findings not only provided a promising drug candidate for clinical treatment of ICH but also shed light on the future drug discovery against ICH.

Sulfate conjugation of daphnetin by the human cytosolic sulfotransferases.

ETHNOPHARMACOLOGICAL RELEVANCE: In Turkey, daphnetin-containing Daphne oleoides is used as a folk medicine for treating rheumatic pain and lumbago. A daphnetin-containing traditional Chinese medicine tablet, named Zushima-Pian, is available in China for treating rheumatoid arthritis. The present study aimed to investigate the metabolism of daphnetin through sulfation in cultured human cells and to identify the human cytosolic sulfotransferase(s) (SULT(s)) that is(are) capable of mediating the sulfation of daphnetin. MATERIALS AND METHODS: Cultured HepG2 human hepatoma cells and Caco-2 human colon carcinoma cells were labeled with [(35)S]sulfate in the presence of different concentrations of daphnetin. Thirteen known human SULTs, previously expressed and purified, as well as cytosols of human kidney, liver, lung, and small intestine, were examined for daphnetin-sulfating activity using an established sulfotransferase assay. RESULTS: [(35)S]sulfated daphnetin was found to be generated and released by HepG2 cells and Caco-2 cells labeled with [(35)S] sulfate in the presence of daphnetin. Among the 13 known human SULTs, SULT1A1, SULT1A2, SULT1A3, SULT1B1, and SULT1C4 displayed significant sulfating activity toward daphnetin. Of the four human organ samples later tested, small intestine and liver cytosols displayed considerably higher daphnetin-sulfating activity than those of lung and kidney. CONCLUSION: The results derived from the present study showed unequivocally that daphnetin could be sulfated in cultured human cells and by purified human SULT enzymes as well as human organ cytosols. The information obtained provided a basis for further studies on the metabolism of daphnetin through sulfation in vivo.

Sulfation of 6-Gingerol by the Human Cytosolic Sulfotransferases: A Systematic Analysis.

Previous studies have demonstrated the presence of the sulfated form of 6-gingerol, a major pharmacologically active component of ginger, in plasma samples of normal human subjects who were administered 6-gingerol. The current study was designed to systematically identify the major human cytosolic sulfotransferase enzyme(s) capable of mediating the sulfation of 6-gingerol. Of the 13 known human cytosolic sulfotransferases examined, six (SULT1A1, SULT1A2, SULT1A3, SULT1B1, SULT1C4, SULT1E1) displayed significant sulfating activity toward 6-gingerol. Kinetic parameters of SULT1A1, SULT1A3, SULT1C4, and SULT1E1 that showed stronger 6-gingerol-sulfating activity were determined. Of the four human organ samples tested, small intestine and liver cytosols displayed considerably higher 6-gingerol-sulfating activity than those of the lung and kidney. Moreover, sulfation of 6-gingerol was shown to occur in HepG2 human hepatoma cells and Caco-2 human colon adenocarcinoma cells under the metabolic setting. Collectively, these results provided useful information relevant to the metabolism of 6-gingerol through sulfation both in vitro and in vivo.

[Effects of Acupuncture on Neurofunction and Neuropsychological Factors of Chronic Alcoholic Peripheral Neuropathy Patients].

OBJECTIVE: To observe the effects of acupuncture on neurofunction and neuropsychological factors of chronic alcoholic peripheral neuropathy (CAPN) patients. METHODS: Totally 120 CAPN patients were assigned to the common treatment group, acupuncture group A, and acupuncture group B according to random digit table, 40 in each group. All patients recieved conventional drug therapy. Besides, patients in the acupuncture group A were additionally needled at Pishu (BL20), Weishu (BL21), Xuehai (SP10), Yinlingquan (SP9), Zusanli (ST36), Yanglingquan (GB34), Jiexi (ST41), Xuanzhong (GB39), Xiangu (ST43),Taixi (KI3), Quchi (LI11), Waiguan (SJ5), Hegu (LI4), and so on. On these bases patients in the acupuncture group B were needled at Sishencong (EX-HN1), Yintang (EX-HN3), Neiguan (PC6), Taichong (LR3), Sanyinjiao (SP6), and Taiyang (EX-HN5). Acupuncture was performed once a day, 14 times as a course; and then once on every other day, 14 times in total for 4 weeks. All treatment lasted for 8 successive weeks. Neuropathy Impairment Score in the Lower Limbs (NIS-LL), Neurological Severity Score (NSS), Hamilton Depression Scale (HAMD), and Hamilton Anxiety Scale (HAMA) were assessed, motor nerve conduction velocity (MCV) and sensory nerve conduction velocity (SCV) were detected before and after treatment. RESULTS: After 8 weeks of treatment the scores of NIS-LL and NSS significantly decreased in the 3 groups, with statistical difference as compared with before treatment (P < 0.05). Scores of NIS-LL and NSS decreased more in acupuncture groups A and B than in the common treatment group (P < 0.05), and more obvious in acupuncture group B (P < 0.05). Compared with the same group before treatment, MCV and SCV of median nerve, ulnar nerve, common peroneal nerve and tibial nerve increased in acupuncture treatment group A and B after 8-week treatment (P < 0.05). MCV of median nerve, MCV and SCV of common peroneal nerve and tibial nerve significantly increased in the common treatment group (P < 0.05). Compared with the common treatment group, SCV of median nerve, MCV and SCV of ulnar nerve, common peroneal nerve and tibial nerve obviously increased in acupuncture treatment groups A and B after treatment (P < 0.05). MCV of ulnar nerve, MCV and SCV of common peroneal nerve and tibial nerve obviously increased more in acupuncture treatment group A than in acupuncture treatment group B (P < 0.05). At week 8 after treatment scores of HAMD and HAMA were obviously lowered in acupuncture groups A and B, with statistical difference as compared with before treatment (P < 0.05). The scores of HAMD were also decreased in the common treatment group, as compared with before treatment (P < 0.05). At week 8 after treatment scores of HAMD and HAMA were obviously lowered more in acupuncture treatment group B than in acupuncture treatment group A (P < 0.05). CONCLUSION: Acupuncture therapy could effectively improve the neurofunction of CAPN patients, and improve complicated anxiety and depression by additionally needling at Sishencong (EX-HN1), Yintang (EX-HN3), Taichong (LR3), Sanyinjiao (SP6), and Taiyang (EX-HN5).

Expression analysis of a novel pyridoxal kinase messenger RNA splice variant, PKL, in oil rape suffering abiotic stress and phytohormones.

Pyridoxal kinase is key enzyme for the biosynthesis of pyridoxal 5'-phosphate, the biologically active form of vitamin B6, in the salvage pathway. A pyridoxal kinase gene, BnPKL (GenBank accession No. DQ463962), was isolated from oilseed rape (Brassica napus L.) following water stress through rapid amplification of complementary DNA (cDNA) ends. The results showed that the gene had two splice variants: PKL and PKL2. PKL, the long cDNA, encodes a 334 amino acid protein with a complete ATP-binding site, pyridoxal kinase-binding site and dimer interface site of a pyridoxal kinase, while PKL2, the short cDNA, lacked a partial domain. Southern blot showed that there were two copies in Brassica napus. The expression of BnPKL cDNA could rescue the mutant phenotype of Escherichia coli defective in pyridoxal kinase. Real-time reverse transcription-polymerase chain reaction revealed that the relative abundance of two transcripts are modulated by development and environmental stresses. Abscisic acid and NaCl were inclined to decrease PKL expression, but H2O2 and cold temperatures induced the PKL expression. In addition, the PKL expression could be transiently induced by jasmonate acid at an early stage, abscisic acid, salicylic acid and jasmonate acid enhanced the PKL expression in roots. Our results demonstrated that BnPKL was a pyridoxal kinase involved in responses to biotic and abiotic stresses.

[The impact of Yishenqinghuo recipe on alveolar bone reconstruction of rats with experimental periodontitis].

PURPOSE: To evaluate the impact of Yishenqinghuo recipe on alveolar bone reconstruction of rats with experimental periodontitis. METHODS: 12-months old Spague-Dawley rats were selected for the study. All rats were randomly divided into 4 groups. Group A: control group (with no periodontitis model, fed with the same dosage of saline as Group D); Group B: model group (with periodontitis model, fed with the same dosage of saline as Group D); Group C: high-dosage group (with periodontitis model, fed with doubling dosage of medicine as Group D); Group D: equivalent-dosage group (with periodontitis model, fed with clinical equivalent effective dosage of medicine). After being gavaged with medicine/saline for 3 months, the alveolar bone were collected to make undecalcified histological slices and evaluate the changes of alveolar bone histomorphometric parameters. All the results were analyzed by ANOVA, with the use of SAS 6.04 software package. RESULTS: Compared with group B, the formation of osteoid were increased and had less free ending trabecular in group C and D. The percent age of trabecular area (%Tb.Ar) and node-terminus ratio (NTR) of alveolar bone for this 4 groups were 75.24+/-3.82/1.49+/-0.12, 45.78+/-6.70/0.48+/-0.08, 73.33+/-4.20/1.33+/-0.06 and 67.69+/-2.83/1.26+/-0.10,respectively. This two parameters of group B were much lower than that in other three groups (P<0.01). At the same time, the NTR in group D was significantly lower than that in group A (P<0.01). The osteoid area (Os.Ar) of different groups were (88.44+/-7.52) microm(2), (145.37+/-13.91) microm(2), (211.10+/-22.96) microm(2) and (201.22+/-24.75) microm(2) (transformation variables), respectively. The Os.Ar of group A was lowest, and that in group B was lower than group C and D, there were significant differences(P<0.01). CONCLUSIONS: From this study, we conclude that Yishenqinghuo recipe is in favor of enhancing alveolar bone quantity, improving bone structure and reconstructing bone loss of rats with experimental periodontitis. Supported by National Key Technologies R&D Program (Grant No.2004BA726) and Natural Science Foundation of Shanghai Municipality (Grant No.03ZR14081).

Genetic diversity and population structure of Lamiophlomis rotata (Lamiaceae), an endemic species of Qinghai-Tibet Plateau.

Lamiophlomis rotata (Lamiaceae), a perennial medicinal herb, is endemic to the Qinghai-Tibet Plateau. A total of 188 individuals from eight natural populations of L. rotata in Qinghai-Tibet Plateau (four from Tibet, two from Yunnan, and two from Qinghai) were analyzed using intersimple sequence repeats (ISSR) and randomly amplified polymorphic DNA (RAPD) techniques. Our results revealed that the level of genetic variation in L. rotata was relatively high (P = 94.85%, I = 0.440 +/- 0.220, H(T) = 0.289 +/- 0.028). Three genetic groups corresponding to the three geographic regions were detected, suggesting significant geographic structure. Our results suggest that the highly structured geographic pattern found in L. rotata might represent diverging evolutionary processes associated with the uplifting of the Qinghai-Tibet Plateau and Quaternary climatic oscillations. These findings imply that as many populations as possible should be preserved in situ for the conservation of this species. Given their genetic variability and peripheral distribution, Qinghai and Yunnan populations should be assigned priority for conservation. Optimal harvesting strategies, domestication and tissue culture of L. rotata should be developed as soon as possible to guarantee its sustainable use.

[Effect of yishenqinghuo recipe on periodontal inflammation and immunity of rats with experimental periodontitis].

PURPOSE: To evaluate the effect of Yishenqinghuo recipe on periodontal inflammation and immunity of rats with experimental periodontitis. METHODS: 12 months old Spague-Dawley rats were used in this study. 78 rats were randomly divided into 4 groups. group A: control group (with no periodontitis, fed with the same dosage of saline as group D); group B: model group (with periodontitis, fed with the same dosage of saline as group D); group C: high dosage group (with periodontitis, fed with double dosage of medicine as group D); group D: equivalent dosage group (with periodontitis, fed with clinical equivalent effective dosage of medicine). After being gavaged with medicine/saline for 3 months, the periodontium was analyzed through histological slices; and the amount of CD4+T,CD8+T, the ratio of CD4+T/CD8+T, IL-2 and IL-1beta in peripheral blood were detected by flow cytometry and ELISA. All the results were analyzed by ANOVA, with the use of SAS 6.04 software package. RESULTS: It was found that the periodontal inflammation of group C and D were improved significantly; the ratio of CD4+T/CD8+T in peripheral blood for this 4 groups were 3.55 +/- 0.94, 2.42 +/- 0.75, 3.23 +/- 1.14 and 3.29 +/- 0.83; the level of IL-2 and IL-1beta were (36.03+/- 2.63/179.04 +/- 17.29) pg/ml, (25.18 +/- 3.08/306.09 +/- 13.38) pg/ml, (38.44 +/- 2.58/176.33 +/- 45.38) pg/ml and (36.81 +/- 2.45/182.13 +/- 43.97) pg/ml. Compared with group B, the ratio of CD4+T/CD8+T for group C and D was significantly rised (P < 1.05); the level of IL-2 increased significantlyand IL-1beta decreased accordingly (P < 0.01). However, there was no significant difference between group C and D (P > 0.05). CONCLUSION: From this study, we conclude that Yishenqinghuo recipe can improve the periodontal inflammation and adjust the immunity of rats with experimental periodontitis. Supported by National "Tenth Five-Year" Key Science and Technology PROject (Grant No. 2004BA720A26) and Natural Science Foundation of Shanghai Municipality (Grant No. 03Zr14081).