hMYH and hMTH1 cooperate for survival in mismatch repair defective T-cell acute lymphoblastic leukemia.

hMTH1 is an 8-oxodGTPase that prevents mis-incorporation of free oxidized nucleotides into genomic DNA. Base excision and mismatch repair pathways also restrict the accumulation of oxidized lesions in DNA by removing the mis-inserted 8-oxo-7,8-dihydro-2'-deoxyguanosines (8-oxodGs). In this study, we aimed to investigate the interplay between hMYH DNA glycosylase and hMTH1 for cancer cell survival by using mismatch repair defective T-cell acute lymphoblastic leukemia (T-ALL) cells. To this end, MYH and MTH1 were silenced individually or simultaneously using small hairpin RNAs. Increased sub-G1 population and apoptotic cells were observed upon concurrent depletion of both enzymes. Elevated cell death was consistent with cleaved caspase 3 accumulation in double knockdown cells. Importantly, overexpression of the nuclear isoform of hMYH could remove the G1 arrest and partially rescue the toxicity observed in hMTH1-depleted cells. In addition, expression profiles of human DNA glycosylases were generated using quantitative reverse transcriptase-PCR in MTH1 and/or MYH knockdown cells. NEIL1 DNA glycosylase, involved in repair of oxidized nucleosides, was found to be significantly downregulated as a cellular response to MTH1-MYH co-suppression. Overall, the results suggest that hMYH and hMTH1 functionally cooperate for effective repair and survival in mismatch repair defective T-ALL Jurkat A3 cells.

Antimicrobial activity of some Alnus species.

OBJECTIVE: The increasing prevalence of resistant microorganisms forced scientists to find new antimicrobial substances from different sources like medicinal plants. The aim of this study was to determine the antimicrobial activities of leaf extracts of some Alnus sp. against some bacteria and a yeast. MATERIALS AND METHODS: Aqueous and ethanolic leaf extracts of A. glutinosa subsp. glutinosa, A. orientalis var. orientalis, A. orientalis var. pubescens were screened for their antimicrobial activities against Staphylococcus aureus ATCC 25923, S. aureus ATCC 43300 (MRSA), Bacillus subtilis ATCC 6633, Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 and Candida albicans ATCC 10231. Broth dilution method was used to determine the antimicrobial activities of plant extracts. RESULTS: Ethanolic extracts of tested species exhibited better antimicrobial activity than aqueous extracts. Ethanolic extracts of tested species possessed activity having MIC values of 0.125-0.250 mg/ml against the tested microorganisms. No antibacterial activity was observed against B. subtilis, E. coli, P. aeruginosa for all the aqueous extracts. Except these aqueous extracts, the others possessed activity having MIC value of 1.000 mg/ml against the tested microorganisms. CONCLUSIONS: To our knowledge, this is the first investigation on the evaluation of antimicrobial activities on aqueous and ethanolic leaf extracts of these species. This study provides significant information about antimicrobial activities of leaf extracts of A. glutinosa subsp. glutinosa, A. orientalis var. orientalis, A. orientalis var. pubescens. It is conceivable that one of the reason for the usage of Alnus glutinosa, in treatment of wound healing in folk medicine, is because of its antimicrobial activity.

Long-term maxillofacial effects of radiotherapy in young nasopharyngeal carcinoma patients: report of 3 cases.

Nasopharyngeal carcinoma (NPC) is a rare and distinct malignancy that arises from the epithelium of the nasopharynx. It accounts almost 1% of all pediatric malignancies. Oral complications of radiotherapy in the head and neck region are the result of the deleterious effects of radiation on salivary glands, oral mucosa, bone, dentition, masticatory musculature, and temporomandibular joints. Here we present 3 male NPC patients 13, 14 and 15 years old. One of them had stage III and the others stage IV diseases. Administered dose of radiation was 66 Gy for case I, 70 Gy for case II and 68 Gy for case III. The follow-up period was more than 12 months except for case III and all of them were disease free in their last visit. All attended dental clinics for dental and TMJ problems. Dentitions were severely affected, trismus and severe xerostomia. Long-term effects of radiotherapy which has a great impact on patients' quality of life and the role of supportive care and minimizing the late effects of ionizing radiation are discussed.

Prilocaine irrigation for pain relief after transurethral resection of the prostate.

PURPOSE: This study aimed to examine the effect of postoperative addition of a local anesthetic agent to irrigation fluid on the alleviation or prevention of pain after transurethral resection of the prostate (TURP). PATIENTS AND METHODS: This prospective, double blind, placebo-controlled study included 50 patients aged 50 to 87 years. The patients were randomized into two groups. As each patient began to experience postoperative pain after spinal anesthesia wore off, 30 mL of 2% prilocaine was injected into the irrigation solution bags, and the bladder was irrigated (prilocaine group). The irrigation solution used in the control group was prilocaine-free. Visual analog scale (VAS) scoring was used to assess the severity of pain. Bladder irrigation in all patients was discontinued 1 day postsurgery. RESULTS: There was not a statistical difference in surgical parameters between the two groups. All patients in the prilocaine group were satisfied with the analgesic efficacy of prilocaine, except for two (8%). The mean number of irrigation solution bags (3000 mL) used for each patient in the prilocaine group was 7.04 ± 1.2. Prilocaine-related side effects were not observed. Conversely, pain developed in all but two patients in the control group. The mean number of irrigation solution bags used for each patient in the control group was 7.6 ± 1.8. Mean VAS pain score was 0.35 ± 0.12 and 5.10 ± 3.26 in the prilocaine and control groups, respectively (P<0.001). CONCLUSION: Prilocaine solution safely alleviated postoperative pain in the patients who underwent TURP. The use of continuous bladder irrigation with a diluted prilocaine solution consistently decreased the need for parenteral analgesics.

Radical quenching activity, ferric-reducing antioxidant power, and ferrous ion-chelating capacity of 16 Ballota species and their total phenol and flavonoid contents.

The ethyl acetate, methanol, and water extracts of 16 Ballota species (Family Lamiaceae)-Ballota acetabulosa, Ballota antalyanse, Ballota cristata, Ballota glandulosissima, Ballota inaequidens, Ballota larendana, Ballota latibracteolata, Ballota macrodonta, Ballota nigra ssp. anatolica, B. nigra ssp. foetida, B. nigra ssp. nigra, B. nigra ssp. uncinata, Ballota pseudodictamnus ssp. lycia, Ballota rotundifolia, Ballota saxatilis ssp. brachyodonta, and B. saxatilis subsp. saxatilis-were screened for their 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical quenching, ferric-reducing antioxidant power, and ferrous ion-chelating capacity at 1mg/mL. Hispanolone, a major diterpene found in the Ballota genus, was also tested in the same manner. Total phenol and flavonoid contents of the extracts were determined by Folin-Ciocalteau and AlCl(3) reagents, respectively. The extracts showed insignificant quenching activity against DPPH radical, but they had moderate antioxidant activity (0.597 ± 0.03 to 1.342 ± 0.01) in the ferric-reducing test compared to chlorogenic acid (the reference compound) (3.618 ± 0.01). All of the extracts (ranging from 65.1 ± 0.64% to 96.3 ± 0.09%) and hispanolone (97.31 ± 0.30%) exerted a remarkable ferrous ion-chelating effect. The highest total phenol (gallic acid equivalent) and flavonoid (quercetin equivalent) contents were found in the ethyl acetate extract of B. glandulosissima (393.7 ± 3.03 and 140.6 ± 1.97 mg/g of extract, respectively). Therefore, Ballota species could be a good source of natural preservatives in foodstuffs.

Prenatal diagnosis of a case with anencephaly-omphalocele-unilateral absent radial ray.

We report here a female fetus with anencephaly, omphalocele and unilateral radial aplasia. The combination of two of these three malformations were reported in a number of patients diagnosed as Gershoni-Barush syndrome, ORR phenotype, VATER association, Schisis association, OEIS complex, Schinzel phocomelia syndrome and Acalvaria but they have not been reported all together in the same patient up to date. We hypothesize that, some or all these syndromes given above may be related etiologically.

Purification and characterization of glucose 6-phosphate dehydrogenase from Lake Van fish (Chalcalburnus tarichiipallas, 1811) liver

Glucose 6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP+ oxidoreductase, EC 1.1.1.49; G6PD) was purified from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver, using a simple and rapid method, and some characteristics of the enzyme were investigated. The purification procedure was composed of two steps: homogenate preparation and 2¡ä, 5¡ä-ADP Sepharose 4B affinity gel chromatography, which took 7¨C8 hours. Thanks to the two consecutive procedures, the enzyme, having specific activity of 38 EU/mg protein, was purified with a yield of 44.39% and 1,310 fold. In order to control the enzyme purification SDS polyacrylamide gel electrophoresis (SDS-PAGE) was done. SDS polyacrylamide gel electrophoresis showed a single band for enzyme. Optimal pH, stable pH, optimal temperature, Km and, Vmax values for NADP+ and glucose 6- phosphate (G6P) were also determined for the enzyme. In addition, molecular weight and subunit molecular weights were found by sodium dodecyl sulfate polyacrilamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography respectively (AU)

Mediante un método simple y rápido, se identifica la enzima glucosa-6-fosfato deshidrogenasa (..) (AU)

Purification and characterization of glucose 6-phosphate dehydrogenase from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver.

Glucose 6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP+ oxidoreductase, EC 1.1.1.49; G6PD) was purified from Lake Van fish (Chalcalburnus tarichii pallas, 1811) liver, using a simple and rapid method, and some characteristics of the enzyme were investigated. The purification procedure was composed of two steps: homogenate preparation and 2', 5'-ADP Sepharose 4B affinity gel chromatography, which took 7-8 hours. Thanks to the two consecutive procedures, the enzyme, having specific activity of 38 EU/mg protein, was purified with a yield of 44.39% and 1310 fold. In order to control the enzyme purification SDS polyacrylamide gel electrophoresis (SDS-PAGE) was done. SDS polyacrylamide gel electrophoresis showed a single band for enzyme. Optimal pH, stable pH, optimal temperature, Km and, Vmax values for NADP+ and glucose 6-phosphate (G6P) were also determined for the enzyme. In addition, molecular weight and subunit molecular weights were found by sodium dodecyl sulfate polyacrilamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography respectively.

HPLC method for the analysis of harmol, harmalol, harmine and harmaline in the seeds of Peganum harmala L.

A simple and sensitive method for separation and determination of harmol, harmalol, harmine and harmaline has been developed and validated. Harmol, harmalol, harmine and harmaline were separated using a Metasil ODS column by isocratic elution with flow rate 1.5 ml/min. The mobile phase composition was Isopropyl alcohol-Acetonitrile-Water-Formic acid (100:100:300:0.3) (v/v/v/v) and pH adjusted 8.6 with triethylamine. Spectrophotometric detection was carried out at 330 nm. The linear range of detection for harmol, harmalol, harmine and harmaline were between 9.375-250, 30.750-246, 31.250-500 and 31.000-248 microg/ml, respectively. The method described was suitable for the determination of harmol, harmalol, harmine and harmaline in the seeds of Peganum harmala L.

A phase II study of paclitaxel and cisplatin combination chemotherapy in recurrent or metastatic head and neck cancer.

The taxanes are the most active new agents for squamous-cell carcinoma of the head and neck (SCCHN) since the discovery of cisplatin. Our aim was to define the therapeutic efficacy and toxicity of paclitaxel and cisplatin combination therapy in patients with recurrent SCCHN. Patients with locally recurrent or metastatic SCCHN were enrolled in the study. Patients were required to be chemotherapy-naive, and should have completed radiation therapy at least 6 weeks prior to enrollment. A World Health Organization (WHO) performance status of less than 3 was required. Paclitaxel (Taxol, Bristol Myers Squibb Company, Princeton, NJ) and cisplatin therapy (PC) consisted of prophylaxis with pheniramine 50 mg i.v., ranitidine 150 mg i.v. and dexamethasone 20 mg i.v. given prior to paclitaxel 175 mg/m2 as a 3-hour i.v. infusion, followed by cisplatin 75 mg/m2 as a 1-hour infusion with an additional 3000 cc of saline for hydration. This treatment was repeated every 3 weeks for a maximum of six cycles. Patients were evaluated for response after the third and sixth cycles, or at the time of clinical progression. Fifty patients were enrolled in the study. The overall response rate was 32% with a 10% complete response rate. Forty-eight patients were assessable for toxicity. A total of 221 cycles of chemotherapy was given and the most common toxicity was myelosuppression; 7.7% of cycles had grade III-IV neutropenia. Severe neuropathy, nephropathy, mucositis, and emesis were uncommon (<10 %). At a median follow-up period of 25 months, the median overall survival was 10 months and the 1-year progression-free and overall survival rates were 16.7% and 35.2%, respectively. We conclude that patients with recurrent SCCHN have a moderate response to combination chemotherapy with cisplatin and paclitaxel. Given this moderate response rate, it is unlikely that this combination (PC) might ultimately prove to be superior to standard treatment regimens in terms of significant survival advantage.

High-dose epirubicin and cisplatin in locally advanced undifferentiated nasopharyngeal carcinoma.

AIM: Undifferentiated nasopharyngeal carcinoma (UNPC) is a chemosensitive tumour; a randomized study evaluating neoadjuvant chemotherapy with bleomycin/epidoxorubicin/cisplatin (BEC) in addition to conventional radiotherapy has resulted in a better disease-free survival in the chemotherapy arm. The bleomycin infusion in the BEC regimen has necessitated hospitalization for the infusion, and resulted in serious pulmonary toxicity. This study has aimed to omit the bleomycin, and test the efficacy and toxicity of cisplatin (C) and a higher dose of epidoxorubicin (EPI) in patients with locally advanced UNPC. METHODS: Seventy-one patients with locally advanced UNPC were treated with three cycles of C 100 mg/m2 day 1, and EPI 100 mg/m2 day 1 every 3 weeks followed by conventional radiotherapy of 70 Gy. RESULTS: Neoadjuvant chemotherapy was well tolerated. There was only 1-week delay in 14.3% of the patients and no dose modification. Grade III-IV neutropenia occurred in 18.9% of the cycles: none of the patients developed neutropenic fever. No patient progressed during chemotherapy, the complete response rate was 26.8% (95% CI = 16.9-38.6) and the partial response rate was 59.1% (95% CI = 46.8-70.7) for an objective response rate of 85.9% (95% CI = 75.6-93.0) at the end of the three cycles of chemotherapy. After the completion of radiotherapy, the complete response rate increased to 81.7% (95% CI = 70.7-89.9) and the objective response increased to 91.5% (95% CI = 82.5-96.8). The median disease-free interval and the median survival have not been reached. The 5-year disease-free and overall survival rates are 53.0% (95% CI = 43.7-62.0) and 57.2% (95% CI = 48.3-65.2), respectively. CONCLUSION: Neoadjuvant C and EPI, easily administered in the outpatient setting, is an effective and well-tolerated regimen in the treatment of locally advanced UNPC.

Extended peptide-based inhibitors efficiently target the proteasome and reveal overlapping specificities of the catalytic beta-subunits.

BACKGROUND: The 26S proteasome is responsible for most cytosolic proteolysis, and is an important protease in major histocompatibility complex class I-mediated antigen presentation. Constitutively expressed proteasomes from mammalian sources possess three distinct catalytically active species, beta1, beta2 and beta5, which are replaced in the gamma-interferon-inducible immunoproteasome by a different set of catalytic subunits, beta1i, beta2i and beta5i, respectively. Based on preferred cleavage of short fluorogenic peptide substrates, activities of the proteasome have been assigned to individual subunits and classified as 'chymotryptic-like' (beta5), 'tryptic-like' (beta2) and 'peptidyl-glutamyl peptide hydrolyzing' (beta1). Studies with protein substrates indicate a far more complicated, less strict cleavage preference. We reasoned that inhibitors of extended size would give insight into the extent of overlapping substrate specificity of the individual activities and subunits. RESULTS: A new class of proteasome inhibitors, considerably extended in comparison with the commonly used fluorescent substrates and peptide-based inhibitors, has been prepared. Application of the safety catch resin allowed the generation of the target compounds using a solid phase protocol. Evaluation of the new compounds revealed a set of highly potent proteasome inhibitors that target all individual active subunits with comparable affinity, unlike the other inhibitors described to date. Modification of the most active compound, adamantane-acetyl-(6-aminohexanoyl)(3)-(leucinyl)(3)-vinyl-(methyl)-sulfone (AdaAhx(3)L(3)VS), itself capable of proteasome inhibition in living cells, afforded a new set of radio- and affinity labels. CONCLUSIONS: N-terminal extension of peptide vinyl sulfones has a profound influence on both their efficiency and selectivity as proteasome inhibitors. Such extensions greatly enhance inhibition and largely obliterate selectivity towards the individual catalytic activities. We conclude that for the interaction with larger substrates, there appears to be less discrimination of different substrate sequences for the catalytic activities than is normally assumed based on the use of small peptide-based substrates and inhibitors. The compounds described here are readily accessible synthetically, and are more potent inhibitors in living cells than their shorter peptide vinyl sulfone counterparts.

Retrograde labeling of primary sensory neurons with fluorescent latex microspheres: a useful tool for long term tagging of neurons.

In this study we have used fluorescent microspheres to retrogradely label primary sensory neurons in dorsal root ganglia (DRGs). Following injection into peripheral nerves, the animals were allowed to survive up to 480 days. Simple profile count indicates that there is a substantial retention of the labeling still after at least 480 days, i.e. about two-thirds of a rat's life span. Moreover, the appearance of the labeling remains quite distinct. Using established markers for axon damage of DRG neurons, we could detect a slight and transient effect of the peripheral nerve injection on the gene expression pattern. It is concluded that fluorescent microspheres represents an attractive means of tagging neurons in experiments covering long time periods.

LC determination of atropine sulfate and scopolamine hydrobromide in pharmaceuticals.

An accurate, simple, reproducible and sensitive method for the determination of atropine sulfate and scopolamine hydrobromide has been developed and validated. Atropine sulfate and scopolamine hydrobromide were separated using a microBondapack C(18) column by isocratic elution with flow rate 1.0 ml/min. The mobile phase composition was methanol, water, formic acid (165:35:1; v/v/v) and pH adjusted 8.3 with triethylamine. The samples were detected at 230 nm using photo-diode array detector. The linear range of detection for atropine sulfate (I) and scopolamine hydrobromide (II) were between 10.38 and 1038 microg/ml with a limit of quantification (LOQ) of 10.38, 10.00 and 1034 microg/ml with an LOQ of 10.00 microg/ml respectively. The linearity, range, peak purity, selectivity, system performance parameters, precision, accuracy, robustness and ruggedness for (I) and (II) were also shown acceptable values.

The rapid quantitative analysis of phenprobamate and acetaminophen by RP-LC and compensation technique.

For the analysis of phenprobamate and acetaminophen in combination, the main analytical methods used were spectrophotometric compensation technique and Vierordt's method with high performance liquid chromatography, used as an analytical reference method. The first procedure for the simultaneous quantitative determination phenprobamate and acetaminophen by high performance liquid chromatographic (HPLC) method was proposed. The method was standardized using a LiChrosorb RP18-5 column, methanol-water-formic acid (120:80:1 v/v), apparent pH 4.25 with triethylamine, as mobil phase and UV detection at 254 nm. The peak area response versus concentration was linear in a concentration range from 4 to 28 microg ml(-1) of phenprobamate and from 4 to 30 microg ml(-1) for acetaminophen. The correlation coefficients were 0.9999 for phenprobamate and 0.9987 for acetaminophen. The second procedure, based on the compensation technique, is presented for the derivative spectrophotometric determination of binary mixtures with overlapping spectra. The proposed methods, which give thoroughly comparable data, are simple and rapid and allow precise and accurate results.

LC method for the analysis of acetylsalicylic acid, caffeine and codeine phosphate in pharmaceutical preparations.

An accurate, simple, reproducible and sensitive method for the determination of acetylsalicylic acid, caffeine and codeine phosphate has been developed and validated. Acetylsalicylic acid, caffeine and codeine phosphate were separated using a microBondapack C(8) column by isocratic elution with flow rate 1.0 ml/min. The mobile phase composition was 125/125/250/0.5 (v/v) isopropyl alcohol, acetonitrile, water and o-phosphoric acid. The samples were detected at 215 nm using photo-diode array detector. The linear range of detection for acetylsalicylic acid, caffeine and codeine phosphate were between 0.40 and 1000, 0.25 and 250, and 0.48 and 96 microg/ml, respectively. The linearity, range, selectivity, system performance parameters, precision, accuracy, and ruggedness for acetylsalicylic acid, caffeine and codeine phosphate were also shown to have acceptable values.

Spectrophotometric resolution of metronidazole and miconazole nitrate in ovules using ratio spectra derivative spectrophotometry and RP-LC.

Metronidazole and miconazole nitrate in ovules was determined by ratio spectra derivative spectrophotometry and by high-performance liquid chromatography (HPLC). The first method depends on ratio spectra first derivative spectrophotometry, by utilizing the linear relationship between substances concentration and ratio spectra first derivative peak amplitude. The ratio first derivative amplitudes at 242.6 [(1)DD(242.6)], 274.2 [(1)DD(274.2))] 261.8 [(1)DD(261.8))] 273.5 [(1)DD(273.5))]and 281.5 [(1)DD(281.5)] nm were selected for the assay of metronidazole and miconazole nitrate, respectively. The second method is based on high-performance liquid chromatography on a reversed-phase column using a mobile phase of methanol-water-phosphoric acid (30:70:0.20 v/v) (pH 2.8) with programmable detection at 220.0 nm. The minimum concentration detectable by HPLC was 0.9 microg ml(-1) for metronidazole and 0.3 microg ml(-1) for miconazole nitrate and by ratio derivative spectrophotometry 4.0 microg ml(-1) for metronidazole and 0.5 microg ml(-1) for miconazole nitrate. The proposed procedures were successfully applied to the simultaneous determination of metronidazole and miconazole nitrate in ovules with a high percentage of recovery, good accuracy and precision.

Serum and tissue c-erb B2, bcl-2, and mutant p53 oncoprotein levels in nasopharyngeal cancer.

To date there are no prognostic factors that can account for the biology and disease behavior in nasopharyngeal cancer. Therefore, identification of new factors that can help in predicting the behavior of the disease and characterizing the subgroup with more aggressive tumors more likely to benefitfrom chemotherapy is important. In this study, c-erb B2, bcl-2, and mutant p53 protein levels were investigated in sera and tumor tissue of patients with nasopharyngeal cancer. Serum c-erb B2 levels were significantly higher in the patients than in the healthy subjects. No meaningful difference was observed between the serum and tissue levels of the mutant p53 protein. Tissue bcl-2 concentrations were considerably high. Our results suggest that serum c-erb B2 levels may aid in identifying a subgroup of patients with a poorer response rate to first-line treatment.

Locally advanced nasopharyngeal carcinoma: computed tomography findings, clinical evaluation, and treatment outcome.

PURPOSE: We present our experience with computed tomography (CT) for delineating the extent of bone erosion in nasopharyngeal carcinoma (NPC) and propose that a new subdivision of Stage T4 disease be added to the staging criteria for cases of minimal bone disease, defined as erosion of the base of the sphenoid or the pterygoid without cranial nerve (CN) involvement. METHODS AND MATERIALS: We retrospectively reviewed the clinical findings, radiological findings, and treatment outcome in 64 patients with Stage T4 NPC, diagnosed according to the American Joint Committee on Cancer 4th edition criteria. The median follow-up was 34 months (range, 3-118 months). Statistical analyses were performed using the chi-square test, the Kaplan-Meier method, and the log-rank test. RESULTS: Local control was achieved in 19 (46%) of 41 patients with CN deficits and 18 (78%) of 23 patients without CN deficits (p = 0.01). Overall 5-year survival with and without CN deficits was 25% and 58%, respectively (p = 0.01). When the 16 patients with minimal bone disease were compared to the remaining 48 patients, there were significant differences in local control rates (87% vs. 48%, p = 0. 006) and 5-year survival rates (68% vs. 28%, p = 0.008). CONCLUSION: Among patients with Stage T4 NPC, a subgroup of patients with only minimal bone disease may have a more favorable prognosis, which may have a considerable bearing on our approach to this patient group.