Deletion of mouse MsrA results in HBO-induced cataract: MsrA repairs mitochondrial cytochrome c.

PURPOSE: Considerable evidence indicates a role for methionine sulfoxide reductase A (MsrA) in lens cell resistance to oxidative stress through its maintenance of mitochondrial function. Correspondingly, increased protein methionine sulfoxide (PMSO) is associated with lens aging and human cataract formation, suggesting that loss of MsrA activity is associated with this disease. Here we tested the hypothesis that loss of MsrA protein repair is associated with cataract formation. To test this hypothesis we examined the effect of MsrA deletion on lens opacity in mice treated with hyperbaric oxygen, identified lens mitochondrial proteins oxidized upon deletion of MsrA and determined the ability of MsrA to repair the identified proteins. METHODS: Wild-type and MsrA knockout mice were treated or not treated with 100 treatments of hyperbaric oxygen (HBO) over an 8 month period and lenses were examined by in vivo light scattering measurements documented by slit-lamp imaging. Co-immunoprecipitation of MsrA was conducted against five specific protein representatives of the five complexes of the electron transport chain in addition to cytochrome c (cyt c). Cyt c in lens protein from the knockout and wild-type lenses was subjected to cyanogen bromide (CNBr) cleavage to identify oxidized methionines. Methionine-specific CNBr cleavage was used to differentiate oxidized and un-oxidized methionines in cyt c in vitro and the ability of MsrA to restore the activity of oxidized cyt c was evaluated. Mass spectrometry analysis of cyt c was used to confirm oxidation and repair by MsrA in vitro. RESULTS: HBO treatment of MsrA knockout mice led to increased light scattering in the lens relative to wild-type mice. MsrA interacted with four of the five complexes of the mitochondrial electron transport chain as well as with cyt c. Cyt c was found to be aggregated and degraded in the knockout lenses consistent with its oxidation. In vitro analysis of oxidized cyt c revealed the presence of two oxidized methionines (met 65 and met 80) that were repairable by MsrA. Repair of the oxidized methionines in cyt c restored the activity of cytochrome c oxidase and reduced cytochrome c peroxidase activity. CONCLUSIONS: These results establish that MsrA deletion causes increased light scattering in mice exposed to HBO and they identify cyt c as oxidized in the knockout lenses. They also establish that MsrA can restore the in vitro activity of cyt c through its repair of PMSO. These results support the hypothesis that MsrA is important for the maintenance of lens transparency and provide evidence that repair of mitochondrial cyt c by MsrA could play an important role in defense of the lens against cataract formation.

The effect of vitamin C or vitamin E supplementation on basal and H2O2-induced DNA damage in human lymphocytes.

There is a wealth of epidemiological information on antioxidants and their possible prevention of disease progression but very little of the research on antioxidants has involved intervention studies. In this study, the potential protective effect of vitamin C or E supplementation in vivo against endogenous and H2O2-induced DNA damage levels in lymphocytes was assessed. The supplementation involved fourteen healthy male and female non-smokers mean age 25-53 (SD 1.82) years, who were asked to supplement an otherwise unchanged diet with 1000 mg vitamin C daily for 42 d or 800 mg vitamin E daily for 42 d. DNA damage in H2O2-treated peripheral blood lymphocytes (PBL) and untreated PBL before and after supplementation, and during a 6-week washout period was assessed using an ELISA. At each sampling time-point, the red cell concentrate activities of superoxide dismutase, catalase and glutathione peroxidase were also determined. Supplementation with vitamin C or vitamin E decreased significantly H2O2-induced DNA damage in PBL, but had no effect on endogenous levels of DNA damage. The activities of the antioxidant enzymes superoxide dismutase and glutathione peroxidase were suppressed during the supplementation period. These supplementation regimens may be used to limit the possible adverse effects of reactive oxygen species (including those produced during the course of an immune response) on lymphocytes in vivo, and so help to maintain their functional capacity.

Activated B cells express increased levels of costimulatory molecules in young autoimmune NZB and (NZB x NZW)F(1) mice.

Polyclonal B cell activation is a hallmark of autoimmune disease in NZB and (NZB x NZW)F(1) (NZB/W) mice. However, the mechanism by which this activated cell subset facilitates disease development is unknown. We recently showed that resting B cells from these mice demonstrate enhanced expression of costimulatory molecules in response to CD40 crosslinking (Jongstra-Bilen et al., J. Immunol. 159,5810-5820, 1997). This led us to question whether activated B cells expressed costimulatory molecules in vivo. Using flow cytometry we found that NZB and NZB/W mice have an increased proportion of splenic B cells expressing B7.1 and elevated levels of B7.2 and ICAM-1. These B cells isolate within the low-density activated population and possess the phenotypic characteristics of marginal zone B cells. The levels of B7.1 on the activated B cell population are similar to those induced by CD40 stimulation raising the possibility that activated B cells in NZB and NZB/W mice provide costimulatory signals to self-reactive T cells leading to loss of tolerance.

Tli1, a resistance locus for carcinogen-induced T-lymphoma.

Within 180 days after injection with N-methyl-N-nitrosourea (MNU), 83.5% of AKR/J mice and 37.5% of BALB/cJ mice developed T-lymphoma. The high tumor incidence was a dominant trait, as 93% of MNU-injected F1 mice developed T-lymphoma. A genome screen of 285 MNU-injected F2 mice identified a locus, designated T-lymphoma Induced 1 or Tli1, in a approximately 10-cM interval on central Chr 1 between D1Mit87 and D1Mit423 with significant linkage to the incidence of MNU-induced T-lymphoma (P = 0.0004). Injection of BALB/cJ.AKR/J-Tli1 congenic mice with MNU confirmed the presence of Tli1 on central Chr 1. Mice homozygous for the BALB/cJ allele (Tli1bb) were over-represented in the tumor-free F2 mice, while the inheritance of parental alleles of Tli1 in tumor-bearing mice was close to expected. This suggests that the Tli1b allele is recessive and suppresses MNU-induced T-lymphoma development in BALB/cJ mice and in Tli1bb F2 mice. Furthermore, the kinetics of lymphoma development in BALB/cJ and the Tli1 congenic mice suggests that Tli1b acts to suppress lymphomas developing late after injection with MNU. Two known genes that map in the identified genomic interval on central Chr 1 are candidates for Tli1:IL10, encoding the lymphokine IL10, and Cmkar4, encoding the chemokine receptor CXCR4.

The B-cell transmembrane protein CD72 binds to and is an in vivo substrate of the protein tyrosine phosphatase SHP-1.

BACKGROUND: Signals from the B-cell antigen receptor (BCR) help to determine B-cell fate, directing either proliferation, differentiation, or growth arrest/apoptosis. The protein tyrosine phosphatase SHP-1 is known to regulate the strength of BCR signaling. Although the B-cell co-receptor CD22 binds SHP-1, B cells in CD22-deficient mice are much less severely affected than those in SHP-1-deficient mice, suggesting that SHP-1 may also regulate B-cell signaling by affecting other signaling molecules. Moreover, direct substrates of SHP-1 have not been identified in any B-cell signaling pathway. RESULTS: We identified the B-cell transmembrane protein CD72 as a new SHP-1 binding protein and as an in vivo substrate of SHP-1 in B cells. We also defined the binding sites for SHP-1 and the adaptor protein Grb2 on CD72. Tyrosine phosphorylation of CD72 correlated strongly with BCR-induced growth arrest/apoptosis in B-cell lines and in primary B cells. Preligation of CD72 attenuated BCR-induced growth arrest/death signals in immature and mature B cells or B-cell lines, whereas preligation of CD22 enhanced BCR-induced growth arrest/apoptosis. CONCLUSIONS: We have identified CD72 as the first clear in vivo substrate of SHP-1 in B cells. Our results suggest that tyrosine-phosphorylated CD72 may transmit signals for BCR-induced apoptosis. By dephosphorylation CD72. SHP-1 may have a positive role in B-cell signaling. These results have potentially important implications for the involvement of CD72 and SHP-1 in B-cell development and autoimmunity.

Lymantria dispar nucleopolyhedrovirus hrf-1 expands the larval host range of Autographa californica nucleopolyhedrovirus.

The gypsy moth (Lymantria dispar) is nonpermissive for Autographa californica nucleopolyhedrovirus (AcNPV) infection. We previously isolated a gene, host range factor 1 (hrf-1), from L. dispar nucleopolyhedrovirus that promotes AcNPV replication in Ld652Y cells, a nonpermissive L. dispar cell line (S. M. Thiem, X. Du, M. E. Quentin, and M. M. Berner, J. Virol. 70:2221-2229, 1996). In the present study, we investigated the ability of hrf-1 to alter the larval host range of AcNPV. Bioassays using recombinant AcNPV bearing hrf-1 were conducted with insect larvae by use of oral infection. AcNPV bearing hrf-1 was infectious for neonate L. dispar larvae, with a 50% lethal concentration of 1.2 x 10(5) polyhedral inclusion bodies/ml of diet, which is similar to that of wild-type AcNPV for permissive hosts. AcNPV can kill neonate L. dispar larvae at high doses, but it does not kill third-instar larvae. However, electron microscopy studies of AcNPV-inoculated third-instar larvae revealed virus replication in the midgut cells. PCR analyses indicated that the virus was AcNPV. These results suggest that the block for AcNPV infection of L. dispar larvae is its inability to spread systematically from primary infection sites in the midgut epithelium and that this barrier is leaky in neonates. hrf-1 allows AcNPV to overcome this barrier. AcNPV recombinants bearing hrf-1 were also significantly more infectious for Helicoverpa zea, a resistant species, suggesting that the blocks for AcNPV infection of L. dispar and H. zea larvae may be similar.

Oxidant-induced stress response in lymphoid cells.

The stress response to reactive oxygen species is an important defence system which can reduce their potential to induce biomolecule damage. In this investigation the effect of exposing Molt-3 lymphoblastoid cells or peripheral blood lymphocytes to a non-toxic dose of hydrogen peroxide (10 microM) was studied. Cellular response to a subsequent high dose of hydrogen peroxide (100-200 microM) was assessed by measurement of growth, viability, proliferation and DNA damage (lymphocytes only) and intracellular activities of the enzymes, superoxide dismutase, glutathione peroxidase and catalase (Molt-3 only). The results indicate that pretreatment of lymphocytes with 10 microM hydrogen peroxide can elicit a response which is protective against DNA damage normally inducible in these cells by subsequent exposure to toxic doses of hydrogen peroxide. It appears from the results with Molt-3 cells that altered activities of glutathione peroxidase may contribute to this enhanced resistance to hydrogen peroxide.

Effects of environmental factors on the in vitro potency of azithromycin.

The effects of media, pH, cations, serum, CO2 or anaerobic atmosphere, inoculum size and time of incubation on the in vitro potency of azithromycin were determined. The potency of azithromycin against all genera was particularly sensitive to changes in pH. The MIC for Staphylococcus aureus strains ranged from 50 micrograms/ml at pH 6 to less than or equal to 0.025 micrograms/ml at pH 8; for erythromycin the MIC change was less (1.6 to 0.05 micrograms/ml). Incubation for 18 h in 5% CO2 or an anaerobic atmosphere (10% CO2, 10% H2, 80% N2) lowered the pH by approximately 0.8 units with gram-negative organisms and 0.4 units with gram-positive organisms. This resulted in an MIC eight times greater than the aerobic MIC. In addition, the MIC100 for azithromycin and erythromycin against Bacteroides strains growing in Wilkins-Chalgren broth fell from 3.1 micrograms/ml in the anaerobic atmosphere to 0.2 and 0.4 micrograms/ml, respectively, when using the Oxyrase enzyme system to remove oxygen. With the Oxyrase system, the pH of the medium at the MIC remained at 7.2, while it fell to 6.7 in the anaerobic gas mixture. An increase in potency for both agents was also observed with other anaerobic species when using the Oxyrase system. The addition of serum produced an increase in potency of azithromycin and erythromycin that correlated with an increase in pH during incubation, despite the use of buffered media. Adding cations to Mueller-Hinton broth resulted in increased MICs for gram-negative organisms; the highest increases observed were four-fold for Escherichia coli.(ABSTRACT TRUNCATED AT 250 WORDS)

Lack of emergence of significant resistance in vitro and in vivo to the new azalide antibiotic azithromycin.

In vitro experiments were performed in which 6 to 12 strains of Staphylococcus aureus, Streptococcus pyogenes, Haemophilus influenzae and Enterobacteriaceae were passaged nine times in sub-lethal concentrations of azithromycin or control antibiotics. Streptococcus pyogenes and Staphylococcus aureus quickly became resistant to rifampin as the MIC90 increased from 0.1 to greater than 50 micrograms/ml for both species. The MIC90 of azithromycin, erythromycin, amoxicillin and cefaclor increased by three dilutions for Staphylococcus aureus. The MIC values of azithromycin for Streptococcus pyogenes, Haemophilus influenzae and Enterobacteriaceae strains did not change significantly. However, for Haemophilus influenzae and the Enterobacteriaceae strains, the MIC values of erythromycin and oral cephalosporins increased four-fold. In the in vivo experiments, mice infected with Staphylococcus aureus or Escherichia coli contaminated sutures were administered azithromycin for three days, and on day 6 viable bacterial cells were recovered from the infection site. The sustained tissue concentrations of azithromycin indicated that the organisms would have been continuously exposed to azithromycin at the site of infection. Colonies isolated from azithromycin-treated and non-treated mice were cultured and their susceptibility to azithromycin compared. The azithromycin MIC values for Staphylococcus aureus cultures from treated and non-treated animals were identical. The azithromycin MICs for Escherichia coli recovered from treated animals were on average, less than one dilution higher than for control cultures. Emergence of significant resistance to azithromycin in the laboratory was not observed with the pathogens tested.

Generation patterns of immunocytochemically identified cholinergic neurons in rat brainstem.

Combined [3H]thymidine autoradiographic and choline acetyltransferase (ChAT)-immunocytochemical techniques were used to answer questions concerning the generation of specific classes and subclasses of cholinergic neurons in rat brainstem. First, the generation of rostrally and caudally located neurons of the same class (i.e. somatic efferent oculomotor and hypoglossal nuclei, respectively) were compared. Results indicated that, although embryonic day 11 (E11) was the peak birthday for both nuclei, hypoglossal neurons were generated significantly earlier than oculomotor neurons, indicating a caudorostral generation gradient for brainstem somatic motor nuclei. Second, the generation patterns of 3 different subclasses of motor neurons at the same brainstem level were compared; namely those of the somatic efferent hypoglossal nucleus (XII), the general visceral efferent dorsal nucleus of the vagus (X), and the predominantly special visceral efferent nucleus ambiguus. All 3 subclasses of cholinergic cells had the same peak day (E11) and overall period of generation (E11-12). However, statistical analyses indicated a precocious generation of nucleus ambiguus, but no developmental differences between N, XII and N. X. It is suggested that nucleus ambiguus is formed earlier than N. XII and N. X, due to its more ventral location within a ventrodorsal neurogenetic gradient. Third, the generation patterns of different classes of large cholinergic neurons were examined. Specifically, the birthdays of cholinergic non-motor projection neurons of the pedunculopontine-laterodorsal tegmental nuclei (PPT-LDT) were contrasted to those of the cholinergic brainstem motor neurons. The peak birthdays of both rostrally and caudally located motor neurons were two days earlier than those of the PPT-LDT neurons. Thus, large cholinergic cells projecting to peripheral targets are born significantly earlier than those projecting within the CNS, even though the former are located more rostrally on the caudorostral neurogenetic gradient. This represents an apparent exception to the emerging rule that cholinergic neurons obey the general gradients of neurogenesis manifest in the regions of the central nervous system where they reside.

Embryonic development of four different subsets of cholinergic neurons in rat cervical spinal cord.

The developmental stage at which a neuron becomes committed to a neurotransmitter phenotype is an important time in its ontogenetic history. The present study examines when choline acetyltransferase (ChAT) is first detected within each of four different subsets of cholinergic neurons previously identified in the cervical enlargement of the spinal cord: namely, motor neurons, partition cells, central canal cluster cells, and dorsal horn neurons. By examining the temporal sequence of embryonic development of these cholinergic neurons, we can infer the relationships between ChAT expression and other important developmental events. ChAT was first detected reliably on embryonic day 13 (E13) by both biochemical and immunocytochemical methods, and it was localized predominantly within motor neurons. A second group of primitive-appearing ChAT-positive cells was detected adjacent to the ventricular zone on E14. These neurons seemed to disperse laterally into the intermediate zone by E15, and, on the basis of their location, were tentatively identified as partition cells. A third group of primitive ChAT-immunoreactive cells was detected on E16, both within and around the ventral half of the ventricular zone. By E17, some members of this "U"-shaped group appeared to have dispersed dorsally and laterally, probably giving rise to dorsal horn neurons as well as dorsal central canal cluster cells. Other members of this group remained near the ventral ventricular zone, most likely differentiating into ventral central canal cluster cells. Combined findings from the present study and a previous investigation of neurogenesis (Phelps et al.: J. Comp. Neurol. 273:459-472, '88), suggest that premitotic precursor cells have not yet acquired the cholinergic phenotype because ChAT is not detectable until after the onset of neuronal generation for each of the respective subsets of cholinergic neurons. However, ChAT is expressed in primitive bipolar neurons located within or adjacent to the germinal epithelium. Transitional stages of embryonic development suggest that these primitive ChAT-positive cells migrate to different locations within the intermediate zone to differentiate into the various subsets of mature cholinergic neurons. Therefore, it seems likely that spinal cholinergic neurons are committed to the cholinergic phenotype at pre- or early migratory stages of their development. Our results also hint that the subsets of cholinergic cells may follow different migration routes. For example, presumptive partition cells may use radial glial processes for guidance, whereas dorsal horn neurons may migrate along nerve fibers of the commissural pathway. Cell-cell interactions along such diverse migratory pathways could play a role in determining the different morphological, and presumably functional, phenotypes expressed by spinal cholinergic neurons.

Fate of mutagenic activity during conventional treatment of municipal wastewater sludge.

The mutagenic activity of wastewater was followed during conventional activated sludge treatment at a municipal plant. Raw wastewater was initially screened for mutagenic potential, using the AmesSalmonella/mammalian microsomal test and employer tester strains. The combined raw wastewater produced dose-related mutagenic responses in the presence of S9 metabolic activation. Raw wastewater from domestic sources alone was not mutagenic. Mutagenic activity was observed throughout the treatment process. Activated sludge prior to chlorination contained the highest specific mutagenic activity. Chlorination decreased the specific mutagenic activity at pH 11. Mutagenic activity in municipal wastewater containing industrial discharges is not removed by conventional treatment processes and can be enhanced by activated sludge treatment.

The v-ski oncogene encodes a truncated set of c-ski coding exons with limited sequence and structural relatedness to v-myc.

The nucleotide sequence of a biologically active v-ski gene from a cloned proviral segment shows that ski is a 1,312-base sequence embedded in the p19 region of the avian leukosis virus gag gene. The v-ski sequence contains a single open translational reading frame that encodes a polypeptide with a molecular mass of 49,000 daltons. The predicted amino acid sequence includes nuclear localization motifs that have been identified in other nuclear oncoproteins. It also contains a proline-rich region and a set of cysteine and histidine residues that could constitute a metal-binding domain. Two regions of the amino acid sequences of v-ski and v-myc are related, and the two proteins exhibit similar distributions of hydrophobic and hydrophilic amino acids. Cloned segments of the chicken c-ski proto-oncogene totaling 65 kilobases have been analyzed, and regions related to v-ski have been sequenced. The results indicate that v-ski is derived from at least five coding exons of c-ski, that it is correctly spliced, and that it is missing c-ski coding sequences at both its 5' and 3' ends. The c-ski and avian leukosis virus sequences that overlap the 5' virus/v-ski junction in Sloan-Kettering virus contain an 18-of-20-base sequence match that presumably played a role in the transduction of ski by facilitating virus/c-ski recombination.

Obesity: is it really a risk factor in thrombophlebitis?

Obesity is assumed to be a risk factor in the occurrence of thrombophlebitis. We studied 168 consecutive patients retrospectively; 33 were men and 135 women, with an average age of 34 (range 27 to 41) years. All patients had a gastric bypass because of obesity, with a minimum of 100 lb over normal weight. The mean weight was 279.2 lb (range 191 to 500). Only three patients had a history of deep vein thrombophlebitis, with no thromboembolism. Eighty-four of the patients were studied preoperatively by noninvasive means (Doppler, impedance plethysmography [IPG], phleborheography [PRG]); 12 had evidence of old disease, and two had a history of treated deep vein thrombophlebitis. No patient had prophylactic therapy. The incidence of clinical deep vein thrombophlebitis was zero; noninvasive evaluation in 64 patients demonstrated no abnormality. Postoperative thromboembolism, which occurred in three of 168 (1.8%) patients, was confirmed by ventilation-perfusion scan and pulmonary angiogram. The mortality from thromboembolism was less than 1% (1/168 patients). Extreme obesity may not necessarily constitute a major risk factor in the occurrence of postoperative deep vein thrombophlebitis and thromboembolism. Prophylactic medications and therapy may add inappropriate risk, undue cost, and unnecessary discomfort, and must be weighed against a mortality of less than 1%.

Organochlorine contamination in shorebirds from Washington state and the significance for their falcon predators.

Arctic-breeding shorebirds collected in western Washington state during winter and spring, and a comparative sample collected in coastal California during the winter were analyzed for organochlorine contaminants to determine the potential impact of these residues on populations of peregrine falcons (Falco peregrinus) and merlins (F. columbarius) which prey upon shorebirds in western Washington. Dunlins (Calidris alpina), an important winter prey for falcons in western Washington, were collected between 1975 and 1981. During winter 1980-81, dunlins carried low organochlorine residues; DDE levels ranged from 0.01 to 1.2 ppm, and PCB levels ranged from 0.02 to 0.82 ppm (wet weight). Levels of other organochlorine contaminants (HCB, Chlordane compounds, Dieldrin, and Heptachlor Epoxide), analyzed in a subsample of dunlins, were consistently lower than DDE and PCB levels, and ranged from 0.001 to 0.22 ppm (wet weight). Dunlins in western Washington did not significantly increase their DDE or PCB burdens over the 1980-81 winter. A decline in DDE residues between 1978 and 1981 was noted, and declines in PCB residues from both 1975 and 1978 to 1980-81 were noted. Residues in other wintering shorebirds from western Washington were similar. Wintering sanderlings (Calidris alba) from California, revealed much higher DDE contamination than in Washington (up to 32 ppm, wet weight). Spring migrant shorebirds in western Washington contained both low and very high DDE residues (up to 417 ppm, wet weight). There is evidence suggesting these high DDE concentrations are accumulated along the Pacific coast of North America.

An improved method of toluidine blue counterstaining of neurons in immunocytochemical preparations.

Immunocytochemical methods can identify individual neurons and processes made immunoreactive by virtue of the antigens they contain. However, frequently it is also useful to visualize surrounding nonimmunoreactive cells, but immunocytochemical procedures often interfere with the quality of subsequent counterstaining. This report describes an improved method of counterstaining immunocytochemical specimens with either aged (at least 1 year) or concentrated solutions of toluidine blue. This technique combines well with immunocytochemical preparations of at least two antigens, i.e., choline acetyltransferase and glutamic acid decarboxylase, to delineate nonimmunoreactive somata. Additionally, a method of photographing these color preparations is described that, by the use of an appropriate filter, allows one to illustrate sections essentially with and without blue counterstain in black and white photomicrographs.

Transforming Sloan-Kettering viruses generated from the cloned v-ski oncogene by in vitro and in vivo recombinations.

The Sloan-Kettering viruses (SKVs) are replication-defective retroviruses that transform avian cells in vitro. Each of the three SKV isolates is a mixture of viruses with genomes ranging in size from 4.1 to 8.9 kilobases (kb) with a predominant genome of 5.7 kb. Using a cDNA representing a sequence, v-ski, that is SKV specific and held in common by the multiple SKV genomes, we generated a restriction map of the 5.7-kb SKV genome and molecularly cloned a ski-containing fragment from SKV proviral DNA. Southern hybridization and sequence analysis showed that the cloned DNA fragment consisted of the 1.3-kb ski sequence embedded in the p19gag sequence and followed by the remaining 5' half of the gag gene and small portions of both the pol and env genes. A large deletion encompassing the 3' half of gag and the 5' 80% of pol was mapped to a position about 1 kb downstream from the 3' ski-gag junction. To determine whether the cloned ski sequence had transforming activity, the ski-containing fragment and a cloned Rous-associated virus 1 (RAV-1) genome were used to construct an analog of the 5.7-kb SKV genome, RAV-SKV. Cotransfection of chicken embryo cells with RAV-SKV and RAV-1 yielded foci of transformed cells whose morphology was identical to that induced by the natural SKVs. The transformed transfected cells produced transforming virus with a 5.7-kb ski-containing genome and synthesized a gag-containing polyprotein of 110 kilodaltons (kDa). Several nonproducer clones of RAV-SKV-transformed cells were analyzed, and most were found to synthesize a 5.7-kb SKV RNA and a 110-kDa polyprotein. One clone was found to contain an 8.9-kb SKV RNA, and this clone synthesized a 125-kDa polyprotein. Since both the 5.7- and 8.9-kb genomes and the 110- and 125-kDa polyproteins had been identified in studies on the natural SKVs, the present results not only demonstrate the transforming activity of these individual SKVs but also suggest mechanisms for their generation.

Use of captopril as early therapy for renal scleroderma: a prospective study.

We conducted a prospective study of captopril therapy in patients with scleroderma and combined hypertension and renal insufficiency. In all seven patients studied during a 1-year period, control of blood pressure was achieved, and in six of the seven, renal function stabilized or improved. The total daily dosage of captopril ranged from 32 to 100 mg, divided into doses taken every 6 to 8 hours. Although one patient had a suspected captopril-induced rash for a short time, none of the other patients had any adverse side effects. Renal biopsies were performed in six patients; in three of them, specimens were obtained both at the beginning and at the end of the study. The initial biopsy specimens showed changes that were similar to those described in other reports. Findings on repeat biopsies were unchanged except for evidence of chronicity. In the six patients with controlled blood pressure and improved or stabilized renal function, the improvement was maintained for 1 1/2 to nearly 3 years on this drug therapy. Using specific measurements of skin compliance and vascular blood flow in the upper extremities, we could detect no evidence, however, of concomitant improvement in these other features of the disease. Although the blood pressure was controlled with captopril, one patient had progressive skin induration, one had progressive pulmonary insufficiency, and another had progressive renal failure.