Transcriptional and metabolic adaptation of human neurons to the mitochondrial toxicant MPP(+).

Assessment of the network of toxicity pathways by Omics technologies and bioinformatic data processing paves the road toward a new toxicology for the twenty-first century. Especially, the upstream network of responses, taking place in toxicant-treated cells before a point of no return is reached, is still little explored. We studied the effects of the model neurotoxicant 1-methyl-4-phenylpyridinium (MPP(+)) by a combined metabolomics (mass spectrometry) and transcriptomics (microarrays and deep sequencing) approach to provide unbiased data on earliest cellular adaptations to stress. Neural precursor cells (LUHMES) were differentiated to homogeneous cultures of fully postmitotic human dopaminergic neurons, and then exposed to the mitochondrial respiratory chain inhibitor MPP(+) (5 µM). At 18-24 h after treatment, intracellular ATP and mitochondrial integrity were still close to control levels, but pronounced transcriptome and metabolome changes were seen. Data on altered glucose flux, depletion of phosphocreatine and oxidative stress (e.g., methionine sulfoxide formation) confirmed the validity of the approach. New findings were related to nuclear paraspeckle depletion, as well as an early activation of branches of the transsulfuration pathway to increase glutathione. Bioinformatic analysis of our data identified the transcription factor ATF-4 as an upstream regulator of early responses. Findings on this signaling pathway and on adaptive increases of glutathione production were confirmed biochemically. Metabolic and transcriptional profiling contributed complementary information on multiple primary and secondary changes that contribute to the cellular response to MPP(+). Thus, combined 'Omics' analysis is a new unbiased approach to unravel earliest metabolic changes, whose balance decides on the final cell fate.

A liver-derived rat epithelial cell line from biliary origin acquires hepatic functions upon sequential exposure to hepatogenic growth factors and cytokines.

Withdrawal of promising drug candidates is often due to the detection of liver toxicity. In particular the parenchymal liver cells or hepatocytes are targeted since they are the major sites of drug transport and of metabolite formation and thus also the place where not only detoxification, but also activation of new chemical (NCE) and biological (NBE) entities may occur. Therefore, primary hepatocyte- based cultures are currently the preferred in vitro model to screen for liver toxicity. However, within a few days, they undergo dedifferentiation with loss of liver-specific functionality, including xenobiotic biotransformation capacity, making them only suitable for short-term applications. A plausible alternative to primary hepatocyte cultures that can be maintained for longer periods of time could be the use of liver-derived epithelial cell lines and their optimized derivatives. Therefore, in the present study, we evaluated the stability and the hepatic differentiation potential of a neonatal liver-derived rat epithelial cell line from biliary origin (rLEC). Undifferentiated rLEC stably express the hepatic progenitor markers CEBPA, FOXA2, GJA1, ONECUT1, KRT18 and KRT19 for at least 15 consecutive passages after cryopreservation. Upon sequential exposure to hepatogenic growth factors and cytokines, rLEC generate functional hepatic progeny, expressing mature hepatic markers including Alb, Ahr, Car, C/ebpα, Cx32, Foxa2, Hnf1α, Hnf1ß and Onecut1. Furthermore, an active polarization is observed for the hepatic drug transporters Oatp4 and Ntcp. rLEC-derived hepatic cells also acquire the ability to store glycogen, express genes encoding for key hepatic enzymes as shown by Affymetrix microarray data, and display stable CYP1A1/2- and CYP2B1/2-dependent activities for several weeks at levels comparable to those observed in cultured primary rat hepatocytes. The acquisition of such a stable and active biotransformation capacity is key for the applicability of liver-based in vitro models for long-term toxicity testing.

Cytosine arabinoside induces ectoderm and inhibits mesoderm expression in human embryonic stem cells during multilineage differentiation.

BACKGROUND AND PURPOSE: Teratogenic substances induce adverse effects during the development of the embryo. Multilineage differentiation of human embryonic stem cells (hESCs) mimics the development of the embryo in vitro. Here, we propose a transcriptomic approach in hESCs for monitoring specific toxic effects of compounds as an alternative to traditional time-consuming and cost-intensive in vivo tests requiring large numbers of animals. This study was undertaken to explore the adverse effects of cytosine arabinoside (Ara-C) on randomly differentiated hESCs. EXPERIMENTAL APPROACH: Human embryonic stem cells were used to investigate the effects of a developmental toxicant Ara-C. Sublethal concentrations of Ara-C were given for two time points, day 7 and day 14 during the differentiation. Gene expression was assessed with microarrays to determine the dysregulated transcripts in presence of Ara-C. KEY RESULTS: Randomly differentiated hESCs were able to generate the multilineage markers. The low concentration of Ara-C (1 nM) induced the ectoderm and inhibited the mesoderm at day 14. The induction of ectodermal markers such as MAP2, TUBB III, PAX6, TH and NESTIN was observed with an inhibition of mesodermal markers such as HAND2, PITX2, GATA5, MYL4, TNNT2, COL1A1 and COL1A2. In addition, no induction of apoptosis was observed. Gene ontology revealed unique dysregulated biological process related to neuronal differentiation and mesoderm development. Pathway analysis showed the axon guidance pathway to be dysregulated. CONCLUSIONS AND IMPLICATIONS: Our results suggest that hESCs in combination with toxicogenomics offer a sensitive in vitro developmental toxicity model as an alternative to traditional animal experiments.

Chemopreventive potential and safety profile of a Curcuma longa extract in women with cervical low-grade squamous intraepithelial neoplasia.

OBJECTIVE: To determine whether Curcuma longa Linn extract, NBFR-03, can arrest low-grade squamous intraepithelial neoplasia (LSIL) a 12 week intervention study was performed. METHODS: Of a total of 1473 women undergoing Pap smear screening, 88 cases had LSIL. Only those with persistent LSIL subsequent to antimicrobial therapy, and willing to follow the protocol (N=21), were included for clinical examination, Pap smears, colposcopy, clinical biochemistry, urinalysis and assessment of serum IL-6, being conducted before and after treatment. Standardised NBFR-03 (0.2gm) capsules were administered, twice daily, for 12 weeks. RESULTS: None progressed to higher grade lesion as assessed by Pap smears and colposcopy. Sixteen cases regressed to atypia, ASCUS or inflammatory pattern; 3 persisted as LSIL, 1 discontinued early because of itching, and 1 did not start. None developed any significant abnormality clinically or biochemically. Micrometry showed a significant reduction in nuclear diameter and nucleocytoplasmic ratio after treatment (p<0.02, and <0.05 respectively). Serum IL-6 levels showed a significant decline (mean 248∓ 156 (SEM) vs 27.7∓ 10.5 (SEM) pg/ ml; p<0.05). CONCLUSION: Use of NBFR-03 for 12 weeks was associated with an arrest or regression of LSIL in Pap smears and colposcopy, with reduction in the circulating IL-6 levels.

Serum IL-6 and micrometry of pap smears in women with cervical low-grade intraepithelial lesions.

OBJECTIVE: To assess serum IL-6 in women with or without low-grade squamous intraepithelial lesions (LSILs) in Pap smears and correlate with the nucleo/cytoplasmic (N/C) ratio. METHODS: Manual micrometry was carried out on Pap smears for N/C ratios: Group A, negative findings (N=15); Group B, inflammation without abnormality (N=14); Group C, LSIL with inflammation (N=13). Serum IL-6 was measured in Groups B and C after treatment of nonviral genital infections. Women with pelvic inflammatory or systemic diseases were excluded. RESULTS: The N/C ratio was significantly higher in Group C vs Group B, both before and after treatment of nonviral infections and also vs group A (p<0.001, Students t test). After treatment for non-viral infections serum IL-6 levels were >50 pg/ml in 5/13 cases of Group C and significantly higher than levels in Group B (p<0.05), correlating positively with the N/C ratio in the 13 cases of LSIL (Pearson's coefficient r=0.659, p<0.05). CONCLUSIONS: High peripheral circulating level of IL-6, despite prior treatment of nonviral infections, was observed in more than one third of women with persistent LSIL in Pap smears, and may serve as an additional biomarker for early cervical neoplasia. Long term follow up is indicated.

Invasive Papillary Breast Carcinoma.

We present the case of a 55-year-old postmenopausal female who presented with complaints of a gradually increasing painless subareolar mass in the left breast of 4 months' duration. Left-sided modified radical mastectomy was performed and the specimen was histopathologically diagnosed as invasive papillary carcinoma. Immunohistochemistry confirmed this diagnosis. All 8 excised axillary lymph nodes were negative for malignant cells. Postoperative chemotherapy was given and for the past 6 months, the patient has maintained a regular follow-up on an outpatient basis. She does not have any evidence of either local or distant recurrence of tumour metastases.

Metaplastic carcinoma with predominant chondrosarcoma of the right breast.

A 45-year-old female presented with complaint of a lump in the right breast for the last 6 months. Excisional biopsy had been performed outside our institution, which was suggestive of a pure mammary chondrosarcoma. Modified radical mastectomy of the right breast was performed and histopathology was suggestive of a pure mammary chondrosarcoma of the right breast. One axillary lymph node was positive for metastasis, the rest of the lymph nodes were free of metastasis. Immunohistochemistry was suggestive of a metaplastic carcinoma with a predominant chondrosarcoma in the right breast. Postoperative radiotherapy was given to the patient with a further plan of chemotherapy.

Pure choriocarcinoma of ovary diagnosed by fine needle aspiration cytology.

Pure ovarian choriocarcinoma is extremely rare and can develop as a germ cell tumor or as a metastasis from uterine or tubal gestational choriocarcinoma or rarely from an ovarian pregnancy. The cytomorphologic findings have been reported previously in different sites. However, this is the first case of pure ovarian choriocarcinoma diagnosed on cytology to the best of our knowledge. The distinction between a gestational and nongestational choriocarcinoma is difficult. A 19-year-old female patient presented with an irregular per-vaginal bleeding and a mass in lower abdomen. Fine needle aspiration cytology smears of the mass were hypocellular and showed large, multinucleated giant cells and malignant mononucleated cells. Background was hemorrhagic. Serum beta hCG level was 3,80,000 mIU/ml. A diagnosis of choriocarcinoma was offered which was later confirmed by histopathology. The diagnosis of choriocarcinoma on fine needle aspiration cytology is based on the presence of large, multinucleated giant cells and malignant mononucleated cells. A high index of suspicion should be maintained and estimation of serum beta hCG plays a key role in supporting the diagnosis.

Sol-gel synthesized semiconducting LaCo0.8Fe0.2O3-based powder for thick film NH3 gas sensor.

Perovskite type LaCo(x)Fe(1-x)O(3) nanoparticles was synthesized by a sol-gel citrate method. The structural, electrical and sensing characteristics of the LaCo(x)Fe(1-x)O(3) system were investigated. The structural characteristics were performed by using X-ray diffraction (XRD) and transmission electron microscopy (TEM) to examine the phase and morphology of the resultant powder. The XRD pattern shows nanocrystalline solid solution of LaCo(x)Fe(1-x)O(3) with perovskite phase. Electrical properties of synthesized nanoparticles are studied by DC conductivity measurement. The sensor shows high response towards ammonia gas in spite of other reducing gases when x=0.8. The effect of 0.3 wt.% Pd-doped LaCo(0.8)Fe(0.2)O(3) on the response and a recovery time was also addressed.

Anaesthetic Management for Cataract Surgery in VACTERL Syndrome Case Report.

SUMMARY: Eight year old girl, weighing 14 kg with VACTERL syndrome V: Vertebral anomalies, A: Anal malformation, C: Cardiovascular defect, TE: Tracheal and esophageal malformation, R: Renal agenesis, L: Limb anomalies. underwent cataract surgery under general anaesthesia. She had multiple congenital anomalies like esophageal atresia, imperforate anus (corrected), single kidney & radial aplasia. Anticipating problems of gastro-esophageal reflux & chronic renal failure, successful management was done.

Extra-abdominal desmoid tumour of the leg.

Extra-abdominal desmoid tumour is a rare tumour and only a few cases occurring in the limbs have been reported. A 35-year-old woman presented with gradually increasing swelling in the upper leg. She had a mild, dull, aching pain in the tumour. Wide local excision was done and the tumour was found mainly in the subcutaneous tissue, which histopathologically proved to be an extra-abdominal desmoid tumour. This case had an abnormal radiological appearance of peripheral calcification of tumour and saucer-shaped lesion in the underlying tibial cortex. The patient had no recurrence at two years follow-up.

Weakness of extensor hallucis longus after removal of non-vascularised fibula as an autograft.

The upper three-quarters of the fibula is commonly used as a non-vascularised autograft. Subsequent to this isolated weakness of extensor hallucis longus may occur. We have studied 26 patients in whom the upper and middle thirds of the fibula had been harvested as a graft through Henry's posterolateral approach. Isolated weakness of extensor hallucis longus was found after operation in ten patients but not in the remainder. EMG and nerve-conduction studies confirmed injury of the nerve to extensor hallucis longus in those with weakness. We dissected 40 cadaver limbs and found that those in which the nerve to extensor hallucis longus ran close to the fibular periosteum were at risk. The injury is mostly incomplete and recovery occurs within four to six months.

Efficacy and safety of the first parenteral selective COX-2 inhibitor, parecoxib sodium, in adult patients with postoperative pain.

Parecoxib, a prodrug of valdecoxib, a selective COX-2 inhibitor, has been recently introduced for the treatment of moderate to severe postoperative pain. This prospective, open, multicentric study enrolled 260 patients undergoing orthopaedic, gynaecological, dental and general surgery. Postoperatively, patients were treated with parecoxib, 40 mg IM/IV. There was a statistically significant decrease in the mean pain intensity score (p<0.05). At the end of 24 hours, 89.6% of total cases had a very good to total relief of pain. The mean duration of analgesia was 19.26 hours and mean time of onset of analgesia was 16.25 minutes ranging from 11-20 minutes. The laboratory values were within normal limits. The drug was well tolerated. There was no report of any hypersensitivity reaction. This study suggests that parecoxib, in a dose of 40 mg IM/IV, is an effective and safe option for the management of postoperative pain.

Effect of closed mitral valvotomy on spirometric pulmonary function tests in mitral stenosis.

The effect of closed mitral valvotomy on the spirometric pulmonary functions was studied in 25 patients with mitral stenosis. The tests were performed before and after operation, the latter at varying intervals (4 to 6 weeks and 8 to 12 months). The preoperative values were considerably low. After 4 to 6 weeks following surgery, further significant reduction in Forced Vital Capacity (FVC) and Forced Expiratory Volume in one second (FEV1) was observed. This was ascribed to the residual healing process and thoracotomy pain. However, Forced expiratory flow rate during mid segment of FVC (FEF25-75%), which reflects obstruction in small airways, did not show any variation. There was improvement in all the above parameters, 8-12 months after surgery. This suggests definite reversibility in the pulmonary functions following valvotomy.

Hypotensive anaesthesia for spine surgery--nitroglycerin vs halothane.

Thirty patients (ASA I or II) requiring spine surgery under general anesthesia were studied. To induce hypotension, halothane 0.5 to 2.5% (n = 15) or nitroglycerin infusion (1-2 micrograms/kg/min) (n = 15) was used. The parameters studied were blood pressure, blood loss, operating time and recovery score. The systolic blood pressure was maintained between 80-100 mmHg during surgery in both the groups. The blood loss with nitroglycerin was significantly less (202 +/- 114 ml) than halothane group (602 +/- 312 ml). All the patients were alert at the end of surgery in the nitroglycerin group (recovery score 9.8 +/- 0.76) as against the halothane group (7.98 +/- 0.9 p < 0.01). Tachycardia or tachyphylaxis was not observed with nitroglycerin. This study suggests that continuous intravenous infusion of nitroglycerin is effective and safe in reducing blood loss and operating time during spine surgery.