Eclampsia in India Through the Decades.

BACKGROUND: Eclampsia is still prevalent in India with high maternal and perinatal mortality. In India, there is no community-based survey of eclampsia. Sporadic reports of study on eclampsia are being published in medical journals. OBJECTIVES: The objective of this study was to find out the incidence of eclampsia and maternal and perinatal mortality in India. METHODS: Data on incidence, maternal mortality and perinatal mortality of eclampsia were collected from articles published in journals and from book of Abstract published during AICOG conferences from 1980 onwards, along with our own data from 1976 to 2014. Data were analyzed yearwise. RESULTS: Reports published from 1976 to 2015 (January-February) reveal that incidence of eclampsia in India ranges from 0.179 to 5 %, the average being 1.5 %. In the period between 1980 and 1989, the average incidence was 0.92 % and the corresponding figure between 2002 and 2010 was 2.15 %, indicating that there is no reduction in incidence of eclampsia in India over the decades. Maternal mortality in 1982 was 14.12 %, and in 2010 it was 2.2-9 %. Maternal mortality has shown a receding tendency, while perinatal mortality is remaining still high as in 1984 it was 45 % and the corresponding figure in 2010 was 24.5-48 %. CONCLUSION: Incidence of eclampsia in India is about 1.5 %. Detail analysis of data from 1980 to 2015 (January-February) shows that there is no reduction in incidence of eclampsia and perinatal mortality rate over the last few decades. Maternal mortality has shown a slight receding trend.

Immunoelectron microscopy on the topographical distribution of the poliovirus receptor.

The topographical distribution of the poliovirus receptor on the cell surface was demonstrated by immunoelectron microscopy using monoclonal antibodies and immunogold markers. The receptor appeared in small clusters, which were randomly distributed over the cell surface and along cellular processes. The distribution pattern of the clusters corresponded to that of absorbed and immunogold-labelled poliovirus particles and suggests a multivalent organization of poliovirus binding sites. Freeze-fracturing and ultrathin sectioning did not reveal any specific ultrastructures within the plasma membrane at labelled receptor areas. Incubation of native cells with anti-receptor antibodies did not remove the receptor molecule from the cell surface nor did it induce ultrastructural alterations within the plasma membrane. The antibody-receptor complexes exhibited lateral mobility within the plasma membrane and were able to aggregate into large immune complexes after incubation with a second ligand.

Activation of endogenous retroviral genomes in Mov strains of mice.

A collection of Mov mouse strains, each carrying in their germ line a Moloney murine leukaemia virus (M-MuLV) proviral genome at a different chromosomal location, was used to study expression of endogenous retroviruses. No M-MuLV-specific RNA was detected in the non-viraemic Mov strains studied, indicating that less than two copies of RNA are transcribed per cell. Virus expression was seen in three viraemic Mov strains. In Mov-3 mice the provirus was activated shortly after birth, whereas proviruses in Mov-9 and Mov-14 were activated at different stages of embryogenesis. The results suggest that the chromosomal position influences proviral expression during development. The first appearance of virus particles was not accompanied by detectable amounts of viral transcripts, suggesting that viraemia is a consequence of provirus activation in a small, as yet unidentified, population of cells, followed by virus spread and infection of susceptible cells.

Transformation of a human poliovirus receptor gene into mouse cells.

The first step in poliovirus replication is binding of virus to a cellular receptor. Mouse L cells, which are resistant to poliovirus infection because they do not bear a poliovirus receptor, were transformed with HeLa cell (human) DNA to poliovirus sensitivity at a frequency of approximately 1 in 50,000 transformants. Monoclonal antibody directed against the HeLa cell poliovirus receptor site was used in rosette assays to identify poliovirus-sensitive L-cell transformants in a background of L-cell tk+ transformants. A cloned cell line, CM-1, was isolated that displayed a surface component recognized by the anti-poliovirus receptor antibody. CM-1 cells were susceptible to infection with all three poliovirus serotypes, and infection could be blocked by the antireceptor antibody. Poliovirus formed plaques in CM-1 and HeLa cells with equal efficiency. CM-1 and HeLa cells produced infectious poliovirus at a similar rate, although yield of virus in CM-1 cells was about 33% less than the yield in HeLa cells. These results suggest that DNA encoding the HeLa cell poliovirus receptor has been introduced into mouse cells, resulting in the expression of the receptor and susceptibility to poliovirus infection.

Involvement of actin filaments in budding of measles virus: studies on cytoskeletons of infected cells.

Cytoskeletons were prepared from measles virus infected HeLa cells to investigate the involvement of cytoskeletal filaments in virus budding at the plasma membrane. The cytoskeletons retained nearly 80% of measles virus hemagglutinin, the major viral polypeptides, including P, NP, and M, and 2 to 12% of the total cell bound infectivity. As demonstrated with platinum- and carbon-shadowed cytoskeletons, all stages of budding, i.e., virus specific strands, stub-like protrusions, and completely rounded virus particles, are associated with actin filaments composing the outer part of the cytoskeletal network. As shown with ultrathin sections of flat embedded extracted cells, actin filaments identified with heavy meromyosin almost exclusively protrude into virus particles with their barbed ends and are in close association with viral nucleocapsids. The data support previous suggestions that actin is involved in virus budding and show that budding itself is possibly the result of a vectorial growth of actin filaments.

Production of a monoclonal antibody against an epitope on HeLa cells that is the functional poliovirus binding site.

Cell lines of primate origin carry receptors on their plasma membrane which are responsible for the specific binding of poliovirus. This paper describes the isolation and characterization of a monoclonal antibody reacting with the plasma membrane of HeLa cells. The antibody (D171) was selected for its protection of HeLa cells against the cytopathic effect of poliovirus type 1. This protection was found to extend to all three viral serotypes, while the replication of five other viruses in HeLa cells was not affected. The 125I-labelled purified antibody did not react with cell lines derived from pig, dog or rodents but bound specifically to all lines of human or primate origin. Immunoglobulin or Fab fragments of D171 prevented the binding of 35S-labelled poliovirus to HeLa cells. Conversely, nearly all binding sites of 125I-labelled D171 immunoglobulins or Fab fragments could be blocked after preincubation of HeLa cells with poliovirus. These results indicate that D171 recognizes the poliovirus receptor site on different susceptible cells and that practically all D171 binding sites are involved in the specific attachment of poliovirus to the plasma membrane. To determine whether the epitope recognized by D171 could be separated from the receptor for poliovirus, human-mouse cell hybrids were prepared and analysed. In all 40 clones tested, the susceptibility to poliovirus correlated with the binding of D171.

Transformation of mononuclear phagocytes in vivo and malignant histiocytosis caused by a novel murine spleen focus-forming virus.

The study of retrovirus-induced leukaemias in mice is a powerful tool for the elucidation of the normal regulation of the haematopoietic system. The acute murine spleen focus-forming viruses (SFFV) can be classified according to the haematopoietic lineage on which they exert their effects in the adult mouse. Here we report a new SFFV isolate, the AF-1 virus, with the novel ability to transform cells of the mononuclear phagocyte lineage. The virus was isolated from sarcomas that were induced on passage of a cloned Friend helper virus (F-MuLV, 643/22F) in newborn BALB/c mice. We have cloned the transforming defective subunit of the AF-1 viral complex in NRK cells and isolated several subclones. Analysis of the proviral genome in two non-producer cell clones reveals that AF-1 virus contains Harvey v-ras-specific sequences (Fig. 1). Thus, AF-1 virus is closely related to Harvey murine sarcoma virus (Ha-MSV), and is, at present, the only tool by which permanent cell lines can be obtained from mononuclear phagocytes in the mouse.

Lack of retrovirus gene expression in somatic cell hybrids of friend cells and teratocarcinoma cells with a teratocarcinoma phenotype.

Two types of hybrids between cells with erythroid phenotype (Friend cells) and teratocarcinoma cells can be distinguished: cell hybrids with an erythroid phenotype, which release or can be induced to release large amounts of Friend spleen focus-forming virus (F-SFFV) on exposure to bromodeoxyuridine and cell hybrids with a teratocarcinoma phenotype, which do not release Friend virus and are not inducible for F-SFFV release. In this paper, we attempted to relate these differences to the expression of F-SFFV and Friend murine leukemia virus (F-MuLV) functions. Teratocarcinoma phenotype hybrids retained F-SFFV-and F-MuLV-related provirus sequences. They did not express F-SFFV- or F-MuLV-related RNA or proteins. The hybrids differentiated to endoderm-like cells on exposure to retinoic acid or hexamethylene-bis -acetamide. These cells, in contrast to the teratocarcinoma phenotype (uninduced) cells expressing SSEA-1-like antigens, did not express SSEA-1-like antigens; they formed typical, prekeratin-staining cytoskeletal structures and could be induced to release mouse interferon. The differentiating cells, but not the uninduced teratocarcinoma hybrids, were infected productively with F-MuLV or the F-MuLV--F-SFFV complex. They, however, did not express endogenous F-SFFV. Endogenous F-SFFV functions could not be rescued by infection with F-MuLV. Induction of teratocarcinoma hybrids with retinoic acid did not activate endogenous F-MuLV or F-SFFV transcription or protein synthesis. These data demonstrated two control mechanisms of Friend virus repression: one which acted trans during formation of the cell hybrids and was maintained only in teratocarcinoma phenotype cells and the other which acted cis and was still operative during induction of endodermal differentiation.

Differentiation and virus expression in BALB/Mo mice: endogenous Moloney leukemia virus is not activated in hematopoietic cells.

The endogenous Moloney leukemia virus (M-MuLV) in the BALB/Mo substrain of mice is activated during the first week after birth. Virus replication occurs in cells of the lymphatic system. Lymphoid cells therefore represent the target cells for virus replication and leukemic transformation. To investigate whether virus activation occurs during lymphoid cell differentiation, hematopoietic stem cells carrying the endogenous Mov-1 genome were transplanted to sublethally irradiated BALB/c mice. Effective colonization of the recipients was demonstrated by Southern DNA hybridization. No activation of the endogenous Mov-1 genome occurred during a 4-month observation period after the transplantation. Thus the first step in development of disease in BALB/Mo mice involves activation of the Mov-1 locus in a nonhematopoietic cell followed by superinfection of lymphatic cells and subsequent virus replication and virus spread.

Chromosomal position and activation of retroviral genomes inserted into the germ line of mice.

The exogenous Moloney leukemia virus (M-MuLV) was inserted into the germ line of mice by exposing embryos to virus at different stages of embryogenesis. Mice derived from exposed embryos were mosaics with respect to integrated virus. Nine new substrains, designated Mov-5 to Mov-13, were derived, each of which carries a single M-MuLV genome at a different chromosomal position in its germ line. Four substrains, Mov-1 to Mov-4, were derived previously. Restriction enzyme analyses demonstrated that, with the exception of Mov-4 and Mov-6 mice, no major rearrangements or deletions have occurred in the integrated proviral genomes. Infectious virus is not activated in the majority of substrains (Mov-4 to Mov-8 and Mov-10 to Mov-12), whereas the other mice develop viremia. A detailed comparison between Mov-1 and Mov-13 mice demonstrated that the time of virus activation is different. Mov-13 mice activate infectious virus during embryogenesis, leading to a distinct pattern of virus expression in all tissues of the adult, but the viral genome in Mov-1 mice is activated only during the first two weeks after birth, leading to virus expression predominantly in lymphatic organs. Together with previous observations, at least four different phenotypes of virus expression-that is, early virus activation during embryogenesis, virus activation after birth, virus activation late in life and no expression of infectious virus at all-can be distinguished among the 13 substrains. Our results suggest that the chromosomal region at which a viral genome is integrated influences its expression during development and differentiation.

Passive immunotherapy prevents expression of endogenous Moloney virus and amplification of proviral DNA in BALB/Mo mice.

BALB/Mo mice carrying the Moloney murine leukemia virus (M-MuLV) as an endogenous virus become viremic soon after birth and develop leukemia at a later age. M-MuLV-specific gene expression and an increase of virus-specific DNA copies in lymphatic target organs are characteristics of the preleukemic phase. Passive immunotherapy of new born BALB/Mo mice with anti-gp70 glycoprotein or anti-M-MuLV serum prevented viremia and delayed significantly the subsequent development of leukemia. Molecular hybridization experiments showed that both virus-specific genome transcription and virus-specific DNA amplification could be completely suppressed by antiserum treatment. Thus virus-specific RNA concentrations in target organs of immunized BALB/Mo mice of 6 months or older were as low as in normal BALB/c mice. This is an age at which untreated BALB/Mo mice have already developed malignant lymphoma. Our experiments demonstrate that treatment with antiserum interferes with the early events of virus expression and thus prevents the subsequent steps leading to leukemia.

Messenger RNA synthesis in synchronized Chinese hamster ovary cells.

Chinese hamster ovary cells were synchronized without inhibitors by mitotic selection and labelled in G1, S or G2 phase by incubation for 90 min with [3H]- OR [14C]uridine. Purified polyribosomes were extracted with phenol and the polyadenylated mRNA prepared by poly(U)-Sepharose chromatography. Poly-adenylated [3H]uridine-labelled mRNA from the G1 phase of the cell cycle was compared by exponential polyacrylamide gel electrophoresis in formamide with [14C] uridine-labelled polyadenylated nRNA from the S or G2 phase. The electrophoretic patterns obtained correspond to the size range expected for mRNA (7-28 S). No prominent differences were detected between mRNAs synthesized in different phases of the cell cycle. From these data we conclude that the major size classes of polyribosomal poly(A)-containing mRNA are synthesized in equal ratios throughout the cell cycle.

Ultrastructural localization of transcription sites and of RNA distribution during the cell cycle of synchronized CHO cells.

The ultrastructural localization of [3H]uridine-labelled RNA synthesized in the course of the cell cycle of synchronized CHO cells is studied using high resolution autoradiography combined with a differential staining for nucleoproteins. It is shown that sites of RNA transcription can already be visualized on the periphery of chromosomes of apparently late metaphase-early anaphase cells with no visible association with the reforming nuclear membrane. In interphase cells they are associated with the border of intranucleolar chromatin or condensed nucleoplasmic chromatin, wherever this localization is made possible by the degree of chromatin dispersion. In cells labeled for 1 or 3 h, the rate of RNA synthesis is higher in S and G2 than in G1. When cells fixed immediately after a [3H]uridine pulse are compared with those post-incubated for 13 h in isotope-free medium, there is a clear difference in intensity of labelling between the nucleus and the cytoplasm. However, the localization pattern of radioactive RNA in the nucleus is similar for all incubation periods as well as for all phases of interphase. The groups of interchromatin granules are generally labeled weakly with radioactivity associated rather with the periphery of their clusters, or remain unlabelled. These results are discussed in the context of other recent findings concerning the distribution of RNA and RNP-structures in the nucleus.

Fragmentation of chromatin with 125I radioactive disintegrations.

The DNA in Chinese hamster cells was labeled first for 3 h with [3H]TdR and then for 3 h with [125I]UdR. Chromatin was extracted, frozen, and stored at -30 degrees C until 1.0 X 10(17) and 1.25 X 10(17) disintegrations/g of labeled DNA occurred for 125I and 3H respectively. Velocity sedimentation of chromatin (DNA with associated chromosomal proteins) in neutral sucrose gradients indicated that the localized energy from the 125I disintegrations, which gave about 1 double-strand break/disintegration plus an additional 1.3 single strand breaks, selectively fragmented the [125I] chromatin into pieces smaller than the [3H] chromatin. In other words, 125I disintegrations caused much more localized damage in the chromatin labeled with 125I than in the chromatin labeled with 3H, and fragments induced in DNA by 125I disintegrations were not held together by the associated chromosomal proteins. Use of this 125I technique for studying chromosomal proteins associated with different regions in the cellular DNA is discussed. For these studies, the number of disintegrations required for fragmenting DNA molecules of different sizes is illustrated.