Fine needle aspiration cytology of a lipoblastoma.

A lipoblastoma, an uncommon tumor of childhood that can be mistaken for a liposarcoma, was preoperatively diagnosed by fine needle aspiration cytology. The characteristic features on the cytologic smears were the presence of immature fat cells in the form of spindle-shaped cells, stellate cells and vacuolated lipoblasts along with lipocytes. The cytologic diagnosis was confirmed by histologic study of the excised tumor.     

Amphiphilic and hydrophilic domains of human sperm membrane antigens recognized by immunoinfertile sera.

Selected sera from married couples with immunological infertility were used to identify antigens on hydrophilic and amphiphilic domains of human sperm membrane. Out of eight sera, six recognized proteins from the hydrophilic as well as amphiphilic regions of the sperm membrane. Sera were either reactive to acrosome or to equator and tail of human sperms in indirect immunofluorescence assay.     

Vaccines for control of fertility.

Major developments in birth control vaccines are on the horizon. The human chorionic gonadotropin (hCG) vaccine has entered phase II clinical trials after successful completion of phase I studies at 5 centers in India and 4 centers abroad. It is the most advanced vaccine of its type in the world. The trials are being conducted on women of proven fertility who are sexually active. The available results indicate the efficiency of the vaccine to prevent pregnancy in women at or above titres of 50 ng/ml. A vaccine inducing antibodies against gonadotropin releasing hormone (GnRH) has been approved in India for trials in postpartum women, to determine whether immunization can help prolong lactational amenorrhea. The GnRH vaccine is also in clinical trial in prostate cancer patients at 2 centers in India and in Austria and the Dominican Republic. The follicle stimulating hormone (FSH) vaccine is about to enter phase I clinical trial after completing experimental and toxicological studies. A vaccine against FSH has been developed for human males employing ovine FSH (oFSH) as an immunogen. oFSH adsorbed on alum induces antibodies reactive with human FSH in bonnet monkeys. Immunization leads to oligospermia with resultant impairment of fertilization potential. No reduction in testosterone levels has been reported. Research is in progress to identify antigens on spermatozoa, which could serve as vaccine candidates. PH-20, a protein located on the inner acrosomal membrane of capacitated sperms, has been reported to have 100% contraceptive efficacy in both sexes of guinea pigs in active immunization studies. cDNA probes of PH-20 cross-react with genomic DNAs of mouse, rat, hamster, and human. The sperm antigen, lactate dehydrogenase C4 (LDH-C4), is a glycolytic enzyme. Active immunization with LDH-C4 suppressed fertility in mice, rabbits, and baboons. SP-10, which is a testis-specific human sperm protein, is also a promising candidate.     

Probing the expression and function of the P2X7 purinoceptor with antibodies raised by genetic immunization

The cytolytic P2X7 purinoceptor is widely expressed on leucocytes and has sparked interest because of its peculiar ability to induce a large nonselective membrane pore following treatment of cells with ecto-ATP. Antibodies raised against synthetic P2X7 peptides generally work well in Western-Blot analyses but fail to recognize the native protein on the cell surface. Genetic immunization is a useful technique to raise antibodies directed against proteins in native conformation. Using this technique we have generated highly specific polyclonal (rabbit) and monoclonal (rat) anti-P2X7 antibodies that readily detect mouse P2X7 on the surface of living cells by immunofluorescence analyses and flow cytometry. Binding of these antibodies to P2X7 is reduced within seconds after treatment of cells with ATP, suggesting that ligand binding induces a conformational shift and/or the shedding of P2X7. By site directed mutagenesis we have mutated three conserved arginine residues (R294A, R307A, R316A) in the extracellular loop of P2X7 near the second transmembrane region. Each of these mutations results in loss of ATP response. FACS and immunoblot analyses reveal that the R294A mutant is expressed at higher levels than wild-type P2X7 in transfected cells, whereas the R307A and R316A mutants are barely detectable because there is no or very little protein synthesis of these constructs. In accord with its resistance to ATP-induced activation the R294A mutant is not down-modulated from the cell surface by ATP-treatment.     

Effect of nucleoside-5'-phosphates on collagen-induced in vitro mineralization

Nucleoside triphosphates (NTPs) at 4-10 microM concentrations were found to inhibit the rates of collagen-induced in vitro mineralization and ion exchange reactions. The sequential removal of the terminal phosphate groups caused a step-wise decrease in their inhibitory potency. The results suggest that NTPs inhibit the rates of ion uptake and exchange reactions at concentrations much lower than their intracellular physiological concentrations. Thus NTPs may be involved in the control of biological mineralization and the tissues which mineralize under physiological conditions develop a system to locally convert NTPs to NDPs and NMPs.     

A low molecular weight substance purified from human placenta inhibits cAMP-dependent protein kinase and activates protein kinase C

We have purified from human placenta a low molecular mass substance that inhibits cAMP-dependent protein kinase and activates protein kinase C. This protein kinase regulator was purified in three steps: (1) homogenizing placentas in chloroform/methanol and extracting the regulator into water; (2) eluting a strong anion exchange high performance liquid chromatography (HPLC) column with a quaternary gradient; and (3) eluting a reversed-phase HPLC column with a binary gradient. The regulator was found to be highly purified by HPLC, thin-layer chromatography (TLC) and laser desorption ionization mass spectrometry with a molecular mass of 703 Daltons by the latter procedure. The physical and biochemical properties of this protein kinase regulator suggest that it is a phospholipid but it did not co-elute by HPLC or by TLC with any of the known phospholipid activators of protein kinase C.