The effect of topical xylometazoline on Eustachian tube function.

BACKGROUND: Topical nasal decongestants are frequently used as part of the medical management of symptoms related to Eustachian tube dysfunction. OBJECTIVE: This study aimed to assess the effect of topical xylometazoline hydrochloride sprayed in the anterior part of the nose on Eustachian tube active and passive opening in healthy ears. METHODS: Active and passive Eustachian tube function was assessed in healthy subjects before and after intranasal administration of xylometazoline spray, using tympanometry, video otoscopy, sonotubometry, tubo-tympano-aerodynamic-graphy and tubomanometry. RESULTS: Resting middle-ear pressures were not significantly different following decongestant application. Eustachian tube opening rate was not significantly different following the intervention, as measured by all function tests used. Sonotubometry data showed a significant increase in the duration of Eustachian tube opening following decongestant application. CONCLUSION: There remains little or no evidence that topical nasal decongestants improve Eustachian tube function. Sonotubometry findings do suggest that further investigation with an obstructive Eustachian tube dysfunction patient cohort is warranted.

A novel gene THSD7A is associated with obesity.

Body mass index (BMI) is a non-invasive measurement of obesity. It is commonly used for assessing adiposity and obesity-related risk prediction. Genetic differences between ethnic groups are important factors, which contribute to the variation in phenotypic effects. India inhabited by the first out-of-Africa human population and the contemporary Indian populations are admixture of two ancestral populations; ancestral north Indians (ANI) and ancestral south Indians (ASI). Although ANI are related to Europeans, ASI are not related to any group outside Indian-subcontinent. Hence, we expect novel genetic loci associated with BMI. In association analysis, we found eight genic SNPs in extreme of distribution (P⩽3.75 × 10(-5)), of which WWOX has already been reported to be associated with obesity-related traits hence excluded from further study. Interestingly, we observed rs1526538, an intronic SNP of THSD7A; a novel gene significantly associated with obesity (P=2.88 × 10(-5), 8.922 × 10(-6) and 2.504 × 10(-9) in discovery, replication and combined stages, respectively). THSD7A is neural N-glycoprotein, which promotes angiogenesis and it is well known that angiogenesis modulates obesity, adipose metabolism and insulin sensitivity, hence our result find a correlation. This information can be used for drug target, early diagnosis of obesity and treatment.

Altersolanol A: a selective cytotoxic anthraquinone from a Phomopsis sp.

UNLABELLED: The cytotoxic compound Altersolanol A, an anthraquinone derivative was isolated from PM0409092 a fungus of Nyctanthes arbor-tristis (family Oleaceae). It was identified as a Phomopsis sp. by DNA amplification and sequencing of the ITS region. The chemical structure of Altersolanol A was elucidated from its physicochemical properties, 2D NMR spectroscopy and other spectroscopic data. The compound has in vitro cytotoxic activity against 34 human cancer cell lines with mean IC50 (IC70) values of 0.005 µg ml(-1) (0.024 µg ml(-1)) respectively. Altersolanol A, a kinase inhibitor, induces cell death by apoptosis through the cleavage by Caspase-3 and -9 and by decreased anti-apoptotic protein expression. There are several previous reports of the anticancer activity of Altersolanol A, but we report here an extensive study using 36 cell lines which gives wider spectrum of results. SIGNIFICANCE AND IMPACT OF THE STUDY: This study confirms the cytotoxic potential of Altersolanol A isolated from the endophyte Phomopsis sp. (PM0409092) of the plant Nyctanthes arbor-tristis. The compound exhibits in vitro cytotoxicity against 34 human cancer cell lines with mean IC50 (IC70) value of 0.005 µg ml(-1) (0.024 µg ml(-1)). This is an in-depth report of Altersolanol A against a panel of 34 human cancer cell lines and extends observations from previous studies indicating that Altersolanol A can be used for the development of chemotherapeutics. Altersolanol A, a kinase inhibitor, induces cell death by apoptosis through the cleavage of Caspase-3 and -9 and by decreased anti-apoptotic protein expression.

Sonography in acute ureteric colic: an experience in Dhulikhel Hospital.

BACKGROUND: Computed tomography is considered as an imaging modality of choice in acute ureteric colic. However due to concerns regarding radiation exposure, sonograms are re-emerging as imaging methods in such situations. OBJECTIVES: To evaluate the role of sonography in detection of calculus in acute ureteric colic. METHODS: Total 384 patients were enrolled. Hydronephrosis was graded as mild, moderate or severe. Calculus was detected as an intraluminal echogenic focus with distal shadowing with twinkling artifact. Number, size and position of the calculi were assessed. Patients were categorized into four groups:I. ureteric colic only II. ureteric colic with hematuria III. ureteric colic with hydronephrosis and IV. ureteric colic with hematuria and hydronephrosis and then the possibility of detection of calculi has been compared among these groups. RESULTS: Out of 384 patients, 254 were found to have calculi ranging between 2.7-27 mm. Nineteen had in the pelvis/ pelviureteric junction, 64 in proximal ureter, 125 in distal ureter, 6 at iliac crossing and 40 at vesicoureteric junction. Two hundred forty one had single and 14 had multiple calculi. Calculus detection is easier in category III and IV patients. The sensitivity and specificity of ultrasonography were 87.98% and 93.07%. Degree of hydronephrosis is strongly correlated with the number of calculi but weakly correlated with the size of the calculus. CONCLUSION: Sonogram can be used in all cases of acute ureteric colic. Hydronephrosis is the most important finding because it paves the way out for the detection of calculus.

Renal impairment among patients with pelvic organ prolapse in a tertiary care center.

BACKGROUND: Pelvic organ prolapse is a multidimensional phenomenon. The manifestations of renal involvement following pelvic organ prolapse ranges from acute to chronic renal failure and may also lead to end stage renal failure. Prolonged duration and its severity in pelvic organ prolapse are responsible for renal impairment. OBJECTIVES: This study will find out the incidence of renal impairment among women with pelvic organ prolapse undergoing surgical management. It will also help to establish the association of renal impairment with degree and duration of pelvic organ prolapse. METHODS: This prospective study includes cases of pelvic organ prolapse who underwent surgical management from the year 2009-2012 in Dhulikhel hospital. Total 140 cases were assessed with age, duration and different symptoms. Clinical examination was done to find out the degree of prolapse and graded according to Baden Walker classification. Urine routine and microscopic examination, renal function test and abdominopelvic sonography were performed routinely. Renal impairment was assessed using hydronephrosis and creatinine clearance. Creatinine clearance was calculated using patient's weight and serum creatinine value by Cockcroft and Gault formula. RESULTS: Among 140 cases, 8 (5.7%) patients had procedentia, 106 (75.7%) had third degree, 22 (15.7%) had second degree, and four (2.8%) patients had first degree uterine prolapse. The mean duration of prolapse was found to be 11.47 years. Total 5 (3.57%) patients had hydronephrosis. Four had moderate and one had severe hydronephrosis. Total 49 (34.1%) patients had moderate to severe renal failure. Forty six (32%) in stage III moderate reduction in creatinine clearance, 2 (1.4%) with severe reduction and 1(0.7%) in end stage renal failure. CONCLUSION: Renal impairment is a common entity among women with pelvic organ prolapse. Both hydronephrosis and degree renal impairment correlates correspondingly with the duration and severity of prolapse.

Role of ultrasound as compared with ERCP in patient with obstructive jaundice.

BACKGROUND: The diagnosis of obstructive jaundice relies on proper history taking, clinical examination, laboratory investigations and different non invasive imaging modalities like Ultrasonography (USG), Cholangio Computed Tomography (CCT), Magnetic resonance Imaging (MRI) with Magnetic Resonance Cholangio Pancreatography (MRCP) and invasive modalities like endoscopic retrograde cholangiography (ERCP) and percutaneous trans hepatic cholangiography (PTC). OBJECTIVE: To compare the role of ultrasound with endoscopic retrograde cholangiography and to determine the major causes of obstructive jaundice in our prospect. METHODS: This was a prospective, analytical study conducted on 88 patients presenting to Department of Radiodiagnosis and Imaging at Dhulikhel Hospital-Kathmandu University hospital from March 2011 to August 2012 with clinical diagnosis of obstructive jaundice. Sonographic evaluation was performed in Siemens acusion x-150 and x-300. The final diagnosis was made by endoscopic retrograde cholangiography and /or surgery and confirmed histopathologically. RESULTS: The most common benign causes of obstructive jaundice were choledocholithiasis (63%), CBD stricture (12.3%), cholangitis (8%) and pancreatitis (6.85%) whereas cholangio carcinoma (6.85%) and carcinoma head of pancreas (4%) comprised of the malignant causes . Ultrasonography had sensitivity of 100% and specificity of 89% in detecting choledocholithiasis. It was found to be 98.78% sensitive and 83.33% specific in cholangiocarcinoma. Similarly in pancreatitis, the sensitivity of ultrasonography was 97.59% and sensitivity was 66.67%. CONCLUSION: Ultrasonography acts as a valuable diagnostic imaging modality in detecting the causes of obstructive jaundice. Due to its easy availability, non invasive nature and cost effectiveness, it can be considered as the first line imaging technique/ tool. ERCP is the invasive imaging tool and can be used for both diagnostic and therapeutic purpose.

Nuchal translucency in normal fetus and its variation with increasing crown rump length (CRL) and gestational age.

BACKGROUND: Nuchal translucency (NT) is the fluid collection behind the fetal neck which can be measured by ultrasound at 11-14 weeks of gestation. Increase in the nuchal translucency thickness is associated with various congenital anomalies. OBJECTIVE: To study the relationship between nuchal translucency thickness, crown rump length and gestational age in normal fetus. METHODS: Prospective analytical study conducted on 211 pregnant women from March 2011 to August 2012. Measurement of Nuchal translucency thickness and crown rump length was performed by ultrasound at 11-14 weeks of gestation. The relationship between nuchal translucency thickness, crown rump length and gestational age was studied by using linear regression analysis. RESULTS: The mean CRL was 63.67+13.48mm (range 41.2-88mm) and mean NT thickness was 1.55+0.35mm (range 0.8-2.7mm), respectively. The median gestational age was 12.9 weeks. The regression equation which shows relation between median NT thickness and CRL was described as follows: expected NT thickness = 0.013CRL+0.725, (R2 = 0.258, p <0.001). There was increase in the incidence of NT thickness more than or equal to 2.5mm; 1.7% in fetus between 12-12.9 weeks of gestation to 15.1% in fetus between 14.0-14.9 weeks. CONCLUSION: Our study offers normative data of NT thickness in normal fetus, which can be used as reference to screen various chromosomal and congenital abnormalities between 11- 14 weeks of gestation. NT thickness increased with increasing CRL and a false positive rate increases with increasing gestational age.

Cyclin-dependent kinase inhibitor, P276-00, inhibits HIF-1α and induces G2/M arrest under hypoxia in prostate cancer cells.

BACKGROUND: Hypoxia-inducible factor-1 (HIF-1) is a master regulator of the transcriptional response to oxygen deprivation and controls genes involved in glycolysis, angiogenesis, migration and invasion. Overexpression of HIF-1α has been demonstrated in many common human cancers. METHODS: Luciferase reporter gene assay under hypoxia and normoxia was used to demonstrate transcriptional inhibition of HIF-1 by P276-00. Detailed studies such as western blotting, reverse-transcriptase-PCR and immunofluorescence were carried out to elucidate its mechanism of action. Cytotoxic potential of P276-00 under normoxia and hypoxia was determined on prostate cancer cells using CCK-8 assay, and cell-cycle analysis was carried out using flow cytometry. Antiangiogenic activity of P276-00 was demonstrated by migration assay and tube-formation assay. Efficacy study of P276-00 was performed in a PC-3 xenograft model. RESULTS: P276-00 inhibits transcriptional activation of HIF-1 under hypoxia. It suppressed hypoxia-mediated nuclear HIF-1α expression, as well as phosphorylation of Akt and 4E-BP1 and abrogated expression of HIF-1-inducible gene viz. vascular endothelial growth factor. Under hypoxia, P276-00 did not exhibit enhanced cytotoxic activity in prostate cancer cells but arrested them in the G2/M phase of the cell cycle. The tubular formation of human umbilical vein endothelial cells and migration of prostate cancer cells were also inhibited by P276-00 in vitro. In addition, it demonstrated significant in vivo efficacy in the PC-3 xenograft model. CONCLUSIONS: Given its low toxicity profile, its demonstrated antitumor activity and its potential to inhibit the HIF-1 pathway, P276-00 should be considered as antiangiogenic chemotherapy for prostate cancer.

Sonographic measurement of fetal pinna length in normal pregnancies.

BACKGROUND: Many studies have emphasized on fetal pinna measurements and morphologic features to use this structure as an additional marker for fetal chromosomal anomaly. OBJECTIVES: To assess relationship between fetal pinna length and gestation age and develop a nomogram. To assess relationship between fetal pinna length and head circumference and biparietal diameter. METHODS: Fetal pinna measurements from the tip of helix to the end of lobe were obtained prospectively in 850 singleton pregnant women between 15 and 40 weeks gestation Normal case was defined as normal sonographic findings during examination and normal infant examination at birth or both. Final study population was 787. The relationship between gestational age in weeks to pinna length in millimeters was analyzed by simple linear regression. Correlation of fetal ear length measurements with gestational age, biparietal diameter and head circumference were also obtained. RESULTS: Linear relationships were found between fetal pinna length and gestational age [Pinna Length (mm)=1.044xGestational age (weeks) -3.857]. A nomogram of normal pinna length was obtained. High correlation was found between pinna length and gestational age (r=0.942; p less than 0.001), pinna length and head circumference (r=0.931; p less than 0.001). Significant correlation was found between pinna length and biparietal diameter (r=0.934; p=0.004) CONCLUSIONS: The results of this study provide a nomogram for fetal pinna. The study also provides relationship and good correlation between pinna length and other biometric measurements.

Mechanism of bubble coalescence induced by surfactant covered antifoam particles.

Mechanism of inter-bubble coalescence by an aqueous fatty alcohol particle suspension antifoam containing a nonionic surfactant has been investigated. By observing visually two colliding air bubbles in a liquid pool in the presence of the antifoam, a four-step mechanism is identified. The role of the surfactant in the antifoam is, for the first time, proposed. A surface tension gradient due to the local surfactant concentration difference enables a surfactant laden hydrophobic particle located on bubble surface to move from the periphery of a liquid film between two colliding air bubbles to their region of contact. Drop volume tensiometry and macroscopic foam column experiments are used to further prove this observation. Subsequently, the particle bridges and dewets the bubbles resulting in film rupture. The rate of drainage of the liquid film depends on the particle hydrophobicity, which necessitates complete surfactant desorption from particle surface. This is corroborated experimentally by Wilhelmy plate tensiometry. In addition, cryo-scanning electron and atomic force microscopy are used to determine the particle shape and the force for its entry into the bubble.

RAPD Analysis for Determination of Components in Herbal Medicine.

In this study, the RAPD (Random Amplified Polymorphic DNA) technique was employed for determination of the components in an Ayurvedic herbal prescription, Rasayana Churna. One-hundred-and-twenty decamer oligonucleotide primers were screened in the RAPD analysis to identify three Ayurvedic medicines, dried stem of Tinospora cordifolia, dried fruit of Emblica officinalis and dried fruit of Tribulus terestris, the Ayurvedic prescription. Primer OPC-6 simultaneously generated three distinct amplicons, each specific to one component. The marker with 600 bp is specific to Tinospora cordifolia; the marker 500 bp is specific to Emblica officinalis and the remaining marker >1000 bp was present in Tribulus terestris. Presence of three herbal medicines was determined when RAPD reaction with OPC-6 was performed. The technique was proved to contribute to the identification of components in Ayurvedic herbal preparation and thus helping to serve as a complementary tool for quality control.

Nonionic block copolymer antifoams.

Aqueous dispersions of alkoxylated alcohol block copolymer (BCP) drops are investigated as antifoams. A model aqueous nonionic surfactant solution of Polysorbate 20 and an industrial white water suspension are used as foaming systems. Visual evidence obtained using a two-bubble technique involving a CCD camera coupled with high magnification lenses clearly revealed the role of BCP droplets in the bubble coalescence process. The enhancement of bubble coalescence decreased as the temperature increased from 25 to 60 degrees C, which is due to the corresponding decrease in the rigidity associated with the weak interfacial structure and reduced viscosity of the BCP drops. The antifoaming efficiency measured in the macroscopic recirculation foam column increased with temperature from about 13 to 26 degrees C (attaining a maximum) and decreased as temperature increased further. Oscillatory thermo-rheometric measurements showed a sudden increase in the storage modulus (G') by several orders of magnitude, indicating gel formation initiated at about 13 degrees C and having a maximum at around 26 degrees C for an aqueous solution of the BCP above a critical concentration of around 20 wt %. Results obtained using small-angle X-ray scattering, micro-differential scanning calorimetry, and proton nuclear magnetic resonance confirmed the existence of ordered gel-like structures. Furthermore, macroscopic tests using a sparged air foam column showed a significant increase in antifoaming efficiency when highly hydrophobic particles are embedded in the BCP drops dispersed in water.

Meckel-Gruber syndrome.

Meckel- Gruber syndrome is a rare lethal, autosomal disorder. It has been linked to chromosome 17. It consists of a triad of occipital meningoencephalocoele, large polycystic kidneys and post-axial polydactyly. Death is mainly due to pulmonary hypoplasia. We report this rare case which presented with many associated defects.

The immunochemistry of sandwich ELISAs--VI. Greater than 90% of monoclonal and 75% of polyclonal anti-fluorescyl capture antibodies (CAbs) are denatured by passive adsorption.

Quantitative data are presented showing that the method most commonly used to immobilize antibodies in microtiter immunoassays functionally inactivates most of the antibodies. These results were collected using five affinity purified polyclonal antibodies (pAbs) and six monoclonal antibodies (mAbs) specific for fluorescein (FLU) as capture antibodies (CAbs). These CAbs were tested for their ability to capture FLU4.2-BSA after immobilization by passive adsorption, the Protein-Avidin-Biotin-Capture (PABC) system or using previously adsorbed anti-globulins. Results indicate that under optimal conditions, < 10% of monoclonal capture antibody equivalents (CAbeqv) and congruent to 22% of polyclonal CAbeqv remain functional after passive adsorption. Immobilization via the PABC system improved the performance of mAbs by more than five-fold but had less than a two-fold effect on pAbs. Many CAbs immobilized using an anti-globulin retained full activity including the ability to bind two molecules of FLU4.2-BSA/molecule of CAb. The latter result is not necessarily a recommendation for the use of anti-globulin immobilization, since the number of functional CAbeqv per well is not significantly greater than that which can be achieved using passive adsorption.

The immunochemistry of sandwich ELISAs--V. The capture antibody performance of polyclonal antibody-enriched fractions prepared by various methods.

Studies compare the performance of antibody-enriched serum fractions prepared by various methods, when adsorbed on polystyrene microtiter wells as capture antibodies (CAbs) and tested against multivalent antigens. The criteria of performance in the RIA used included antigen capture capacity (AgCC) and the nmol of functional capture sites per microtiter well (CAbt). Affinity purified polyclonal (pAb) and monoclonal antibodies (mAb) were employed as reference CAbs. AgCC was highest for enriched fractions prepared using caprylic acid and a high-pressure SpG affinity column. The performance of capture antibodies is expressed by an equation which was empirically derived and experimentally tested; CAbt x AgCC/ng adsorbed IgG. In terms of this parameter, CAb-enriched fractions prepared with caprylic acid performed best. The data reported also provide insight into solid-phase ligand immunochemistry. Adsorbed polyclonal CAb performed with remarkable homogeneity in percent bound and in Scatchard plots. Values obtained for CAbt from Steward-Petty plots were directly correlated with the length of the LBR of log-log percent bound plots but indicated that less than 10% of the potential capture sites of polyclonal CAbs remained functional after adsorption; mAb showed a more serious loss of activity. The loss of CAbt was a general phenomenon for all preparations tested although relative to their antibody content, certain antibody-enriched fractions retained a higher proportion of CAbt than their affinity-purified counterparts. Comparative studies in which the activity of adsorbed mAb and pAb was compared to the same antibodies immobilized by a non-adsorptive procedure, indicated that adsorbed CAbs also express lower affinity. The studies we report offer a single parameter criterium for comparatively evaluating CAb performance while simultaneously revealing the need to develop immobilization procedures that can preserve CAbt and antibody affinity so that immunoassays with wide dynamic ranges and high AgCC can be developed without waste of antibody.

The physical and functional behavior of capture antibodies adsorbed on polystyrene.

Six monoclonal and two polyclonal antibodies to fluorescein (FLU) were affinity purified and immobilized on Immulon 2 polystyrene as capture antibodies (CAbs): (a) by passive adsorption at pH 9.6, (b) via a streptavidin bridge to a biotinylated carrier molecule, and (c) via an antiglobulin which had been previously adsorbed passively to the polystyrene. Data show that less than 3.0% of the binding sites of monoclonal CAbs and approximately 5-10% of those of polyclonal CAbs were capable of capturing antigen (FLU4.2-BSA) after passive adsorption. Immobilization of CAbs via an antiglobulin or a streptavidin bridge, resulted in the preservation of antibody binding sites to greater than 70% for some monoclonals although immobilization via the streptavidin bridge resulted in the highest number of functional sites/well. The data presented are consistent with studies on other adsorbed proteins which demonstrate that passive adsorption on polystyrene results in the loss of protein function. Furthermore, these data show that generally less than half of the binding sites of antibodies available in solution are available after solid-phase immobilization even when non-adsorptive methods are employed. Some polyclonal anti-FLU also have lower average avidity following passive adsorption compared with CAbs immobilization via a streptavidin bridge. Immunochemical studies revealed that adsorbed polyclonal-CAbs performed like monoclonals when tested with multivalent antigens (FLU10-IgA) but in an expected heterogeneous manner in Scatchard plots when tested using univalent FLU-insulin. This observation implied cross-linking of immobilized CAbs by the multivalent antigen. Because only 5-10% of adsorbed polyclonal CAbs are active, the survivors must be non-randomly distributed in clusters to explain the cross-linking. This was confirmed by scanning electron microscopy which gave rise to the hypothesis that antibodies which retain activity after adsorption, are those present in clusters, i.e., the functional adsorbed CAb is an antibody cluster. Data presented in this report on the behavior of adsorbed CAbs, and reviewed from the work of others for various adsorbed proteins, indicate that the method of passive adsorption at pH 9.6, which is widely used in popular microtiter ELISAs, and which has in many ways revolutionized immunoassay, is a method of protein denaturation. Assayists that utilize passive adsorption of proteins on hydrophobic supports as part of their research need to be cognizant of this phenomenon, while inventors of immunoassay should develop alternative methods of immobilization which do not destroy 90% of the functional activity of solid-phase reactant.

Humoral immunity in root caries in an elderly population. 1.

IgA, IgG and IgM antibody activity (ELISA Units/ml) to Streptococcus mutans, Actinomyces viscous and Escherichia coli CF8 in serum, parotid saliva and whole saliva was measured using the amplified ELISA (a-ELISA) while the concentration (microgram/ml) of each isotype of immunoglobulin as well as albumin and lactoferrin, was determined using sandwich ELISAs. Selection of suitable reagents from those commercially available was based on specificity tests using purified human immunoglobulin; most polyclonal reagents required further absorption to attain class specificity. Cross-absorption studies indicated the absence of patient antibodies that were cross-reactive among the bacteria studied, except for IgM in some cases. Expression of response in ELISA Units (E.U.) per microgram of immunoglobulin, i.e. specific activity, revealed that IgG specific activity was significantly higher in parotid saliva than in either whole saliva or serum for all bacteria studied; serum and whole saliva did not differ except for the higher specific activity in whole saliva to E. coli. The value of one E.U. was determined using the Comparative Antibody-immunoglobulin Capture Assay (CACA). Using this novel method, we estimated that about 0.05 percent of serum IgA was specific for Streptococcus mutans, 0.008 for Actinomyces viscosus and 0.004 for Escherichia coli CF8. The percentage of specific IgM antibodies was higher than for IgA and IgG. The concentration of IgA anti-Streptococcus mutans, Actinomyces viscosus and Escherichia coli levels are approximately 92 ng/ml, 25 ng/ml and 16 ng/ml in whole saliva and 46 ng/ml, 9.4 ng/ml and 6.3 ng/ml in parotid saliva.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiparametric evaluation of retinoic acid-induced terminal differentiation of blastoid cells from acute non-lymphocytic leukemia patients in vitro.

Recent findings that retinoic acid (RA) induces terminal granulocytic differentiation of the human promyelocytic leukemia cell line HL-60 in vitro and blast cell maturation in patients suffering from acute non-lymphocytic leukemia (ANLL) prompted an investigation on the ability of this agent to induce terminal maturation in blast cells from ANLL patients in vitro. We tested the ability of RA at 3 x 10(-6) M, 3 x 10(-7) M and 3 x 10(8-) M concentrations to induce differentiation in blastoid cells from 16 patients with ANLL using cytochemical and cytologic parameters, in addition to cytofluorometric methods. Leukemic cells in primary culture from all the patients underwent cytochemical and biochemical changes after treatment with RA. However, the extent of differentiation-positive cell clones (D+ clones) varied from patient to patient. Morphologic maturation was observed in a significant number of bone marrow samples. Leukocyte alkaline phosphatase and NBT reduction ability of cells, which are biochemical markers of granulocytic differentiation, were also significantly increased with a simultaneous decrease in DNA and RNA synthesis (which was estimated using a Phywe ICP-11 impulse flow cytometer).

Terminal differentiation of human leukemic blasts in response to 12-0-tetradecanoyl-13-phorbol acetate (TPA).

Phorbol ester, 12-0-Tetradecanoyl-13-Phorbol-Acetate (TPA), induces a terminal macrophage-like differentiation of cells from human acute myelogenous leukemia cell lines. We report here that blastoid cells obtained from acute nonlymphocytic leukemia (M1-M2) undergo differentiation-related changes characteristic of macrophage lineage after exposure to TPA. Blast cells from a patient with ANLL-M1-M2 underwent morphological, functional and histochemical changes after treatment with 1 x 10(-7) and 1 x 10(-8) M TPA. The changes included adhesion to the plastic substrate, 2-4 fold increase in the number of NBT positive cells and an increase in the number of alpha-naphthyl-acetate esterase (alpha-NAE) positive cells. These differentiation changes after treatment with TPA were followed by decrease in proliferative index and G1 cells containing high RNA as estimated by flow cytometry. Of the thirteen cases of undifferentiated or unclassified leukemias studied, two failed to respond to TPA. These data suggest that leukemic blasts retain their ability to express a variety of differentiated functions on induction by TPA. Our data gives evidence suggesting that the "switch" into the differentiation pathways occurred after inhibition of proliferation and reduction in the percentage of G1 high RNA containing cells.