[Efficient method for analyzing absorbed ingredients of traditional Chinese medicine genitana macrophyllae radix].

In present study, a method for analyzing the absorbed ingredients of traditional Chinese medicine QinJiao has been developed. A rat everted gut sac (EGS) model has been established, and the transporting capacity of gut sacs was identified by histological examinations. The ingredients including loganic acid, sweroside, gentiopicroside, and swertiamarian in serosal solution absorbed by active transport of rat everted ileum and jejunum from Qinjiao extraction were determined using an HPLC method. Histological integrality of the gut sacs remains perfect and the active transport activity of them is normal within 45 min of the experiment. The HPLC method employed in this study presents high specificity and good correlation. The relative standard deviation of precision of this method is less than 5.5%. Extraction recovery of samples is more than 90%. And stability of the samples in room temperature is perfect. Eight ingredients of Qinjiao absorbed in serosal solution are identified. Furthermore, concentration of Qinjiao extraction significantly affects accumulated absorption and absorption coefficient of the ingredients. However, there is no significant impact on the accumulated absorption and absorption coefficient by diverse of everted gut sections. From above, the EGS techniques might be an efficient method, which can be employed for investigation of absorbed ingredients of Traditional Chinese Medicines.

Angle Stable Interlocking Intramedullary Nails for Tibial Plateau Fractures.

OBJECTIVE: Angle stable interlocking intramedullary nail (ASIN), a novel technique, has rarely been used for treatment of tibial plateau fractures (TPF). This retrospective study was designed to introduce this novel technique, ASIN, as well as to describe the initial experience and verify the effectiveness when ASIN was used for the management for TPF. METHODS: A cohort of 19 cases with closed TPF aged from 18-70 years with at least 23 months follow-up from November 2008 to September 2013 was analyzed retrospectively. All patients underwent the ASIN procedure, which was performed by the same group of surgeons. Perioperative and postoperative parameters like the measurement of radiographic pictures, surgical data, and clinical function were recorded including the changes in treatment. A modified Hohl-Luck radiological and functional score combined with the Hospital for Special Surgery (HSS) score were applied to evaluate the final results and to provide reliable data through the whole procedure when applying the ASIN procedure. RESULTS: The patients were followed up regularly for an average of 26.3 (range, 23-34) months. All patients achieved a bony union at an average of 15.1 weeks with no incidences of malunion, nonunion, or infection. Anatomical reduction of the articular surface was obtained in 16 patients. No secondary failure of fixation occurred. The mean postoperative knee flexion was 122.9°. The modified Hohl-Luck radiological and functional score was excellent and good, respectively, in 16 patients. The mean HSS score was 89.4. CONCLUSION: The angle stable interlocking intramedullary nail system turned out to be a viable alternative protocol in the treatment of tibia plateau fractures and provided satisfactory results, with good fracture reduction, biomechanical fixation, low rates of complications, and passable postoperative knee function.

Toxic effects of Fusarium mycotoxin butenolide on rat myocardium and primary culture of cardiac myocytes.

Mycotoxin toxicosis has been implicated in the etiopathogenesis of Keshan disease (KD), an endemic cardiomyopathy prevailing in some regions of China. Butenolide (4-acetamido-4-hydroxy-2-butenoic acid gamma-lactone, CAS No. 16275-44-8), a mycotoxin produced by several Fusarium species such as Fusarium tricinctum and Fusarium graminearum, is frequently detected from the cereals in the endemic areas of KD. The present study is undertaken to investigate whether this mycotoxin can induce myocardial damage. Exposure of primary culture of cardiac myocytes to butenolide resulted in significant cytotoxicity, manifested by changes in cell morphology and decreases in cell viability. Consistent with the in vitro findings, distinct myocardial toxicity in vivo was observed after administration of rats by gavage with butenolide (10 and 20 mg/kg/day) for 2 months, and the myocardial injuries were characterized by focal necrosis of myocardium and fragmentation of myofiber. Butenolide also induced significant oxidative damage to the myocardium in vitro evidenced by a concentration-dependent lipid peroxidation in the myocardial homogenates, whereas antioxidants superoxide dismutase (SOD), N-acetylcysteine (NAC) and glutathione (GSH) provided significant protections against this oxidative effect. Taken together, these results clearly reveal that butenolide possesses the potential to induce myocardial toxicity. The present findings may reinforce the hypothesis that toxicosis by mycotoxins is one of the etiological factors for KD.

Direct effects of lead (Pb2+) on the relaxation of in vitro cultured rat aorta to acetylcholine.

Lead (Pb(2+)) exposure is related to increased blood pressure or hypertension of human or animals. Abnormal vascular relaxant responses of low level Pb(2+) exposed animals were reported by several studies. However, it is difficult to tell whether these effects were induced directly by Pb(2+) or not. In this study we hypothesized that Pb(2+) can directly affect the relaxation of vessels. Male Wistar rat aortae were removed and cultured in PMRI 1640 with 1 ppm Pb(2+) (4.8 microM lead acetate) for 0.5, 6, 12 and 24h, and then their responses to acetylcholine (ACh) and sodium nitroprusside (SNP) were examined. After incubated for 24h, the relaxation induced by ACh was significantly decreased in Pb(2+) exposed aortic rings. However, there was not significant difference in relaxation induced by SNP between Pb(2+) exposed and control group. The nitrite in the culture media of aortic rings cultured for 24h, measured with Griess method, was significantly decreased in the Pb(2+) exposed group. The expression of endothelial NOS (eNOS) and isoform NOS (iNOS) in the homogenate of aortic rings cultured for 24h was measured by Western blot. The expression of eNOS of the Pb(2+) exposed group was significantly upregulated compared with that of the control group. However, there was no significant difference in the expression of iNOS in control and Pb(2+) exposed group. In conclusion, Pb(2+) was able to directly affect the relaxation of rat aorta. This effect may have some relation with the lower level of NO in the media, though the expression of eNOS was upregulated.

Metallothionein protects against doxorubicin-induced cardiomyopathy through inhibition of superoxide generation and related nitrosative impairment.

Metallothionein (MT) has been shown to be an effective protector against DOX-induced cardiomyopathy, however the involved precise mechanisms are still unknown. The present study was undertaken to clarify whether the inhibition of superoxide generation and related nitrosative damage were involved in the metallothionein attenuation of DOX-induced cardiac injury. MT-I/II null (MT-/-) mice and corresponding wild-type mice (MT+/+) were pretreated with either saline or zinc (300 micromol/kg, s.c., once a day for 2 days) prior to a single dose of DOX (15 mg/kg, i.p.) or equal volume of saline. Animals were sacrificed on the 4th day after DOX administration and samples were collected for further analyses. DOX caused remarkable cardiac damage in both MT+/+ and MT-/- mice as demonstrated by biochemical and histopathological alterations. Zinc pretreatment significantly increased the cardiac MT levels and therefore inhibited the cardiac toxic effects of DOX only in MT+/+ mice, but not in MT-/- mice. Furthermore, elevated formation of superoxide and peroxynitrite were obviously observed after DOX treatment, while these elevation were prevented by MT induction by zinc in MT+/+ mice, but not in MT-/- mice. These findings suggest that metallothionein induction by zinc exhibits protective effects on the cardiac toxicology of DOX, which might be mediated through the prevention of superoxide generation and related nitrosative impairment.

A simple method for the formalin fixation of lungs in toxicological pathology studies.

Optimized lung preparation for detailed structural evaluation is required to improve consistency in preclinical safety evaluation, differences of opinion exist among regulatory agency personnel regarding the optimal methods for routine formalin fixation of lungs from rodent toxicology studies. The simple tracheal ligation fixation method emphasizes tracheal ligation before opening the thorax instead of attempting to re-inflate after lung collapse when opening the thorax. Photomicrographs of this method demonstrated an unprecedented ability to maintain the natural lung architecture, in contrast to the unavoidable changes in the alveolar environment by the intratracheal instillation and vascular perfusion methods. In addition, a comparison of fixation methods on lung morphology in a rodent model of LPS-induced acute lung injury demonstrated that the tracheal ligation fixation method may provide a standard approach for morphometry. Additionally, a TUNEL assay was used to determine the degree of autolysis, which revealed that the autolysis was insignificant in the central areas of each lobe of the lung compared to the lung periphery by tracheal ligation fixation. In conclusion, our novel modified method, which avoids the disadvantages of generating artifacts, fulfills the requirement of preserving the clear, natural morphology of the lung making it suitable and worthy of recommendation for toxicological studies in a good laboratory practice (GLP) lab.

Subchronic toxicological study of two artemisinin derivatives in dogs.

The objective of our study was to profile and compare the systematic changes between orally administered artesunate and intramuscularly injected artemether at a low dose over a 3-month period (92 consecutive days) in dogs. Intramuscular administration of 6 mg kg-1 artemether induced a decreased red blood cell (RBC) count (anemia), concurrent extramedullary hematopoiesis in the spleen and inhibition of erythropoiesis in the bone marrow. We also observed a prolonged QT interval and neuropathic changes in the central nervous system, which demonstrated the cortex and motor neuron vulnerability, but no behavioral changes. Following treatment with artesunate, we observed a decreased heart rate, which was most likely due to cardiac conduction system damage, as well as a deceased RBC count, extramedullary hematopoiesis in the spleen and inhibition of erythropoiesis in the bone marrow. However, in contrast to treatment with artemether, neurotoxicity was not observed following treatment with artesunate. In addition, ultra-structural examination by transmission electron microscopy showed mitochondrial damage following treatment with artesunate. These findings demonstrated the spectrum of toxic changes that result upon treatment with artesunate and artemether and show that the prolonged administration of low doses of these derivatives result in diverse toxicity profiles.

Apoptosis in microencapsulated juvenile rabbit chondrocytes induced by ofloxacin: role played by beta(1)-integrin receptor.

Quinolone(s) (QNs) is widely used in infection therapy due to its good antimicrobial characteristics. However, QNs-induced arthropathy of immature animals has led to restrictions on the therapeutic use of these antimicrobial agents. The exact mechanism(s) of QNs-induced chondrotoxicity remain unknown. In the present study, we investigated the possible mechanism of ofloxacin (one typical QNs)-induced injuries of chondrocytes. Juvenile rabbit joint chondrocytes cultured in alginate microspheres were incubated with ofloxacin at concentrations of 0, 2, 5, 10, 20, and 40 microg/ml for up to 96 h. Concentration of 10 microg/ml ofloxacin induced apoptosis of chondrocyte with visible apoptotic signs, including degradation of poly(ADP-ribose) polymerase, caspase-3 activation, and DNA ladder formation. Furthermore, extracellular signal-regulated kinase 1/2 (phospho-ERK1/2) and growth factor receptor-bound protein 2 (Grb2) were significantly reduced, and similar changes were also observed in the beta(1)-integrin receptor as assessed by immunoblotting. However, the mRNA level of beta(1)-integrin obtained from reverse transcription-polymerase chain reaction remained unchanged. Results of beta(1)-integrin immunoprecipitation have also shown that beta(1)-integrin did not interact with activated intracellular signaling proteins. In addition, ofloxacin did not induce apoptosis and decrease beta(1)-integrin expression in chondrocytes supplemented with Mg(2+), and the ofloxacin-induced apoptosis was caspase-8-dependent, inhibition of which did not affect the expression mode of phospho-ERK1/2 and beta(1)-integrin. Our results demonstrate that ofloxacin affects beta(1)-integrin receptor functions and the ERK mitogen-activated protein kinase signaling pathway, causing caspase-8-dependent apoptosis after exposure of 48 h.