Relationship between baseline severity of depression and antidepressant treatment outcome.

INTRODUCTION: Assessment of depression severity is of key importance, since several clinical guidelines recommend choice of treatment dependent on the depression severity grade. Using different tools to assess baseline severity may result in different outcomes. METHODS: This paper describes the results of a multicentre, naturalistic study investigating the relationship between depression symptom severity (using 4 different measures of symptom severity) and clinical outcome among patients hospitalised for depression (N=1 014). Moreover, the impact of differences between methods of measuring depression severity has been investigated. Statistical analyses (univariate measurements, logistic regression models) were conducted to detect coherences and differences between the various methods of severity categorisation. RESULTS: Results revealed different associations between outcome and classification methods. Response or remission rates varied if baseline severity was assessed by different instruments. Moreover, the number of responders increased with higher baseline severity grades of depression, whereas the number of remitters decreased. Additional analyses dependent on outcome criteria using continuous instead of categorical data revealed similar results. DISCUSSION: Baseline severity may be only one of many other important clinical variables that mediate clinical outcome, but it is surely an important one deserving further research and consideration.

Construction of large DNA segments in Escherichia coli.

Recombinant DNA clones containing large pieces of DNA are useful in the study of large genetic units, but these are difficult to make in most bacterial cloning vectors. A strategy is described that uses general and site-specific recombination to construct large pieces of eukaryotic DNA from smaller cloned segments. The large clones are propagated on F factor-based plasmids in Escherichia coli. They can be easily modified to introduce mutations or rearrangements. These techniques were applied to the construction of large DNA segments from the bithorax complex of Drosophila.

Enhancer point mutation results in a homeotic transformation in Drosophila.

In Drosophila, the misexpression or altered activity of genes from the bithorax complex results in homeotic transformations. One of these genes, abd-A, normally specifies the identity of the second through fourth abdominal segments (A2 to A4). In the dominant Hyperabdominal mutations (Hab), portions of the third thoracic segment (T3) are transformed toward A2 as the result of ectopic abd-A expression. Sequence analysis and deoxyribonuclease I footprinting demonstrate that the misexpression of abd-A in two independent Hab mutations results from the same single base change in a binding site for the gap gene Krüppel protein. These results establish that the spatial limits of the homeotic genes are directly regulated by gap gene products.

Molecular Genetics of the Bithorax Complex in Drosophila melanogaster.

The bithorax complex in Drosophila melanogaster is a cluster of homeotic genes that specify developmental pathways for many of the body segments of the fly. The DNA of the bithorax complex has been isolated, and a region of 195,000 base pairs that covers the left half of the complex is described here. The lesions associated with many of the bithorax complex mutants have been identified, and most are due to DNA rearrangements. Most of the spontaneous mutants have insertions of a particular mobile element named "gypsy." This element affects the functions of sequences removed from the site of insertion. Mutant lesions for a given phenotypic class are distributed over large DNA distances of up to 73,000 base pairs.

Polycomb group repression reduces DNA accessibility.

The Polycomb group proteins are responsible for long-term repression of a number of genes in Drosophila melanogaster, including the homeotic genes of the bithorax complex. The Polycomb protein is thought to alter the chromatin structure of its target genes, but there has been little direct evidence for this model. In this study, the chromatin structure of the bithorax complex was probed with three separate assays for DNA accessibility: (i) activation of polymerase II (Pol II) transcription by Gal4, (ii) transcription by the bacteriophage T7 RNA polymerase (T7RNAP), and (iii) FLP-mediated site-specific recombination. All three processes are restricted or blocked in Polycomb-repressed segments. In contrast, control test sites outside of the bithorax complex permitted Gal4, T7RNAP, and FLP activities throughout the embryo. Several P insertions in the bithorax complex were tested, providing evidence that the Polycomb-induced effect is widespread over target genes. This accessibility effect is similar to that seen for SIR silencing in Saccharomyces cerevisiae. In contrast to SIR silencing, however, episomes excised from Polycomb-repressed chromosomal sites do not show an altered superhelix density.

Chromosomal walking and jumping to isolate DNA from the Ace and rosy loci and the bithorax complex in Drosophila melanogaster.

A chromosomal walk is described that covers 315 X 10(3) base-pairs of DNA from the 87DE region of the third chromosome of Drosophila melanogaster. The walk includes the DNA for the rosy and Ace loci, which code for xanthine dehydrogenase and acetylcholinesterase, respectively. Several dispersed repetitive elements were encountered in the walk. In every case, their positions in the chromosome differed in different strains, and so they are all presumed to be transposable elements. Several rearrangement breakpoints have been localized within the walk, including the break for In(3R) Cbx+R1 (87E1, 2-89E1, 2). One breakpoint fusion fragment of this inversion was isolated to jump from 87E into the cluster of homeotic genes of the bithorax complex, at 89E1-4.

Molecular mapping of genetic and chromomeric units in Drosophila melanogaster.

We have used a set of overlapping cloned segments defining a 315 kb (X 10(3) base-pairs) region of Drosophila melanogaster chromosomal DNA to map the sequences associated with the polytene band-interbands (chromomeric units) and with the lethal complementation groups contained within this region. The molecular map positions of the 13 +/- 1 chromomeric units from the 87D5-6 to 87E5, 6 region of the third chromosome were determined by in situ hybridization of selected segments to the polytene chromosomes. The length of the largest chromomeric unit within the 315 kb region is approximately 160 kb, while that for the smallest is less than 7 kb and may be as short as 3 kb. By mapping the breakpoints of deletions within the 315 kb region, we have located its 12 lethal complementation groups, which include the genes coding for acetylcholinesterase (Ace) and xanthine dehydrogenase (rosy). Comparison of the two molecular maps indicates a one-to-one topographical correlation between the genetic and chromomeric units.

Gene conversion in Drosophila and the effects of the meiotic mutants mei-9 and mei-218.

Simple meiotic gene conversion tracts produced in wild-type females were compared with those from two meiotic mutants, mei-9 and mei-218. The positions and lengths of conversion tracts were determined by denaturing gradient gels and DNA sequencing. Conversion tracts in wild type averaged 885 base pairs in length, were continuous, and displayed no obvious hot spots of initiation. Some unusual conversion events were found in the mei-218 and mei-9 samples, although most events were indistinguishable from wild-type tracts in their length and continuity.

Enhancer traps in the Drosophila bithorax complex mark parasegmental domains.

Eight P elements carrying a beta-galactosidase (lacZ) reporter have been mapped to sites within the Drosophila bithorax complex. The bithorax complex contains three homeotic genes, and at least nine regulatory regions which control their expression in successive parasegments of the fly. The enhancer traps inserted at the promoter of one of the genes, Ultrabithorax, express lacZ in patterns which mimic the Ultrabithorax protein pattern. Enhancer traps in the regulatory regions do not mimic the endogenous genes, but express lacZ globally in the relevant parasegments. Some P elements carry large DNA fragments upstream of the lacZ promoter but internal to the P element. In cases where these internal sequences specify a lacZ pattern, that pattern is generally suppressed when the element is inserted in the bithorax complex. In embryos mutant for genes of the Polycomb group, the lacZ expression from the enhancer traps spreads to all segments. Thus, the enhancer traps reveal parasegmental domains that are maintained by Polycomb-mediated repression. Such domains may be realized by parasegmental differences in chromatin structure.

Comparison of germline mosaics of genes in the Polycomb group of Drosophila melanogaster.

The genes of the Polycomb group (PcG) repress the genes of the bithorax and Antennapedia complexes, among others. To observe a null phenotype for a PcG gene, one must remove its maternal as well as zygotic contribution to the embryo. Five members of the PcG group are compared here: Enhancer of Polycomb [E(Pc)], Additional sex combs (Asx), Posterior sex combs (Psc), Suppressor of zeste 2 [Su (z) 2] and Polycomblike (Pcl). The yeast recombinase (FLP) system was used to induce mitotic recombination in the maternal germline. Mutant embryos were analyzed by staining with antibodies against six target genes of the PcG. The loss of the maternal component leads to enhanced homeotic phenotypes and to unique patterns of misexpression. E(Pc) and Su(z) 2 mutations had only subtle effects on the target genes, even when the maternal contributions were removed. Asx and Pcl mutants show derepression of the targets only in specific cell types. Psc shows unusual effects on two of the targets, Ultrabithorax and abdominal-A. These results show that the PcG genes do not act only in a common complex or pathway; they must have some independent functions.

Studies on the mechanism of heterochromatic position effect at the rosy locus of Drosophila melanogaster.

Experiments are described that extend the characterization of position effect variants of the rosy locus and test possible mechanisms of heterochromatic position effect. Rosy position effect variants exhibit a variegated phenotype with respect to xanthine dehydrogenase activity in malpighian tubules. The breakpoints of the position effect mutations are located on the DNA map of the rosy region outside of the rosy locus DNA; ryps11136 is located in the DNA of the l(3)S12 gene immediately proximal to rosy, whereas ryps1149 is located some 15 kb distal to rosy in the pic locus. Southern blot experiments are described that test and reject the notion that heterochromatic position effect results from underreplication of the position-affected gene. Rather, the results of Northern blots serve to direct attention to position effect as a defect in transcription. Histone region deletion heterozygosity and butyrate-feeding experiments failed to exhibit specific suppression of position effect at the rosy locus.

Molecular mapping of the rosy locus in Drosophila melanogaster.

The DNA from the chromosomal region of the Drosophila rosy locus has been examined in 83 rosy mutant strains. Several spontaneous and radiation-induced alleles were associated with insertions and deletions, respectively. The lesions are clustered in a 4-kb region. Some of the alleles identified on the DNA map have been located on the genetic map by fine-structure recombination experiments. The genetic and molecular maps are collinear, and the alignment identifies the DNA location of the rosy control region. A rosy RNA of 4.5 kb has been identified; its 5' end lies in or near the control region.

Mapping point mutations in the Drosophila rosy locus using denaturing gradient gel blots.

Mutations within the rosy locus of Drosophila were mapped using blots of genomic DNA fragments separated on denaturing gradient gels. DNA sequence differences between otherwise identical small rosy DNA fragments were detected among the mutants as mobility shifts on the blots. Mutations were mapped to within a few hundred base pairs of rosy sequence in 100 of 130 mutants tested--a 77% detection rate. The sequence changes in 43 rosy mutations are presented; all but six of these were single base changes. Thirty-four of 36 sequenced mutations induced by the alkylating agents N-ethyl-N-nitrosourea and ethyl methanesulfonate were transitions. All of the mutations mapped in the rosy transcription unit. Twenty-three of the 43 sequenced mutations change the predicted rosy gene polypeptide sequence; the remainder would interrupt protein translation (17), or disrupt mRNA processing (3).

Molecular analysis of recombination events in Drosophila.

The locations of crossover junctions and gene conversion tracts, isolated in the rosy gene of Drosophila melanogaster, were determined using DNA sequencing and denaturing gradient gel electrophoresis. Frequent DNA sequence polymorphisms between the parental genes served as unselected genetic markers. All conversion tracts were continuous, and half of the reciprocal crossover events had conversion tracts at the crossover junction. These experiments have also identified the sequence polymorphisms responsible for altered gene expression in two naturally occurring rosy variants.

Interactions of Drosophila Ultrabithorax regulatory regions with native and foreign promoters.

The Ultrabithorax (Ubx) gene of the Drosophila bithorax complex is required to specify parasegments 5 and 6. Two P-element "enhancer traps" have been recovered within the locus that contain the bacterial lacZ gene under the control of the P-element promoter. The P insertion that is closer to the Ubx promoter expresses lacZ in a pattern similar to that of the normal Ubx gene, but also in parasegment 4 during embryonic development. Two deletions have been recovered that remove the normal Ubx promoter plus several kilobases on either side, but retain the lacZ reporter gene. The lacZ patterns from the deletion derivatives closely match the normal pattern of Ubx expression in late embryos and imaginal discs. The lacZ genes in the deletion derivatives are also negatively regulated by Ubx and activated in trans by Contrabithorax mutations, again like the normal Ubx gene. Thus, the deleted regions, including several kilobases around the Ubx promoter, are not required for long range interactions with Ubx regulatory regions. The deletion derivatives also stimulate transvection, a pairing-dependent interaction with the Ubx promoter on the homologous chromosome.

Mutations affecting expression of the rosy locus in Drosophila melanogaster.

The rosy locus in Drosophila melanogaster codes for the enzyme xanthine dehydrogenase (XDH). Previous studies defined a "control element" near the 5' end of the gene, where variant sites affected the amount of rosy mRNA and protein produced. We have determined the DNA sequence of this region from both genomic and cDNA clones, and from the ry+10 underproducer strain. This variant strain had many sequence differences, so that the site of the regulatory change could not be fixed. A mutagenesis was also undertaken to isolate new regulatory mutations. We induced 376 new mutations with 1-ethyl-1-nitrosourea (ENU) and screened them to isolate those that reduced the amount of XDH protein produced, but did not change the properties of the enzyme. Genetic mapping was used to find mutations located near the 5' end of the gene. DNA from each of seven mutants was cloned and sequenced through the 5' region. Mutant base changes were identified in all seven; they appear to affect splicing and translation of the rosy mRNA. In a related study (T. P. Keith et al. 1987), the genomic and cDNA sequences are extended through the 3' end of the gene; the combined sequences define the processing pattern of the rosy transcript and predict the amino acid sequence of XDH.

Mechanism of deterioration in renal function in patients with renovascular hypertension treated with enalapril.

Azotemia produced by converting enzyme inhibition in renovascular hypertension was studied in six patients by measurement of glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). Patients with unilateral renovascular hypertension lowered GFR acutely to 40% of control values at 4 hours, but after 4 days it only returned to 75% of control even though ERPF exceeded control values. Calculated filtration fraction (GFR/ERPF) decreased from 0.31 to 0.19 in 4 hours (p less than 0.05) and remained at 0.20 after 4 days of treatment despite increases in GFR and ERPF. In bilateral renovascular hypertension, neither the decreased GFR nor lowered ERPF induced by enalapril showed any tendency to return toward normal with continued treatment. These data are consistent with selective glomerular efferent arteriolar dilation in response to enalapril and suggest that angiotensin converting-enzyme inhibition interferes with the autoregulatory capacity of the kidney in the presence of severe renovascular hypertension.

The effect of 17beta-estradiol-DNA adducts on the replication of exon # 5 of the human suppressor gene p53.

Using a PCR technique, exon # 5 of the human tumor suppressor gene p53 was amplified and ligated into the pCRII vector and transformed into Escherichia coli INV alphaF' competent cells. The cloned exon # 5 was 184 bp long. Evidence is presented to show that after dimethyldioxirane epoxidation, 17beta-estradiol was able to form 17beta-estradiol-DNA adducts and to strongly inhibit the replication of the cloned exon # 5 producing smaller sizes of DNA fragments and introducing errors of incorporation at the 3'-end of the terminating DNAs. The errors occurred mainly at the clusters of the complementary 'G' and 'A' bases on the template strand DNA, presumably, the major sites where the 17beta-estradiol-DNA adducts were formed.

Interstitial nephritis, proteinuria, and renal failure caused by nonsteroidal anti-inflammatory drugs. Immunologic characterization of the inflammatory infiltrate.

Nine patients with the unusual combination of renal failure, nephrotic-range proteinuria, and biopsy-proved interstitial nephritis are described. Six of these patients had received nonsteroidal anti-inflammatory agents (three fenoprofen, one ibuprofen, one zomepirac, and one tolmetin). The remaining three patients had no history of exposure to drugs known to cause interstitial nephritis. Immunologic characterization of the infiltrating cells with monoclonal antibodies showed that the majority of cells in most cases were cytotoxic T cells, although some B cells were present in all cases. Giant collecting duct cells were seen in half the patients with drug exposure but in none of the others. Otherwise, there were no conspicuous morphologic differences between patients with and without drug exposure. Many of the patients had associated glomerular abnormalities. Only the zomepirac and tolmetin recipients showed pure interstitial disease. The three fenoprofen recipients and the zomepirac and tolmetin recipients regained normal renal function after the drug was discontinued. The combination of renal failure, nephrotic range proteinuria, and interstitial nephritis is one form of nephrotoxicity observed in patients treated with nonsteroidal anti-inflammatory agents. However, this lesion, which may be mediated by cytotoxic T cells, may also be seen rarely in patients with no apparent drug exposure.

Acyclovir in mouse cytomegalovirus infections.

A virus-specified thymidine kinase appears to be a general requirement for herpes virus susceptibility to the antiviral effect of acyclovir. Surprisingly, mouse cytomegalovirus (MCMV), which does not encode for a thymidine kinase, is exquisitely sensitive to the drug both in vitro and in vivo. The drug is active against the virus in the absence of a cellular thymidine kinase and the antiviral activity is not diminished in the presence of excess thymidine or a variety of nucleosides and deoxynucleosides. Thus, a thymidine phosphorylation pathway is not required for the drug's activation of this infection. The enzyme system responsible for phosphorylation of the drug has not been identified. Mouse cytomegalovirus mutants resistant to the drug have been isolated, indicating that the antiMCMV effect results from selective inhibition of viral replication rather than indirectly through toxicity to the host cell. Eight resistant mutants appear to be in the same complementation group and seven of the mutants demonstrate coresistance to phosphonoacetic acid, a marker for the DNA polymerase locus of herpes viruses. The evidence to date indicates that the MCMV DNA polymerases is the final site of action of the drug. Investigations of the antiMCMV activity of acyclovir should provide insights into the antiviral effects of this drug and other nucleoside analogs in other herpes virus infections in which the virus does not code for a thymidine kinase (for example, human cytomegalovirus and Epstein-Barr virus).