Examination of Induced Sputum In the Diagnosis of Pneumocystis Carinii Pneumonia

The results of the examination of sputum induced by the inhalation of nebulized hypertonic saline in the diagnosis of Pneumocystis carinii pneumonia (PCP) are presented. In suspected cases of PCP in patients who were either HIV antibody positive or were receiving immunosuppressive therapy, 46 induced sputum specimens were stained using both Grocott's modified Gomori methenamine silver nitrate (GMS) and immunofluorescence staining. In 12 specimens P. carinii cysts were detected by both methods, in four specimens by GMS staining only and in five specimens by immunofluorescence only. The sensitivity of induced sputum examination in the detection of P. carinii cysts was increased by using both of these staining methods on each sputum specimen and the need for more invasive methods of diagnosis was reduced.     

地塞米松抑制卡氏肺孢子菌肺炎β2-防御素和Dectin-1受体的表达

目的研究卡氏肺孢子菌肺炎（Pneumocystis carinii pneumonia,PCP）鼠肺Dectin-1和β2-防御素的表达变化,探讨地塞米松对Deetin-1和B2-防御素的影响与疾病发生的相互关系。方法实验分4组：正常对照组、Pc刺激组、PCP模型组以及PCP模型恢复组。免疫抑制方法建立PCP动物模型,改良四胺银（Groeoti’s methenamine—silver nitrate method,GMS）染色检测Pc包囊;肺组织切片HE染色观察肺组织病理变化;实时荧光定量和Westernblot检测Deetin-1和B2-防御素的mRNA以及蛋白的表达。结果Pc刺激组的Dectin．1和β2-防御素的mRNA以及蛋白的表达明显高于正常对照组（P〈0．05）;Pc刺激组和PCP恢复组的Dectin-1和β2-防御素的mRNA以及蛋白显著高于PCP组（P〈0．05）,而PCP恢复组与PCP组肺部炎症无明显差别。结论对免疫功能正常宿主,Dectin-1受体和p2-防御素可能在防Pc感染中起重要作用;地塞米松抑制了鼠肺Deetin-1和β2-防御素的表达,这可能与PCP疾病的发生和发展有关。     

Plasminogen activator production in a rat model of Pneumocystis carinii pneumonia

Several studies have indicated that the serine protease urokinase-plasminogen-activator (uPA) is an important factor in host defense against pulmonary pathogens. To gain a better insight into the role of uPA in Pneumocystis carinii (P. carinii) pneumonia (PCP), we evaluated PA production in alveolar macrophages (AMs) obtained from rats with steroid-induced PCP. Treatment with cortisone acetate favored PCP in 91% of rats. In the bronchoalveolar lavage (BAL) samples of immunosuppressed rats both with and without PCP, we observed a decrease in uPA activity as well as a decrease in cell number. Urokinase-PA production by AMs was reduced in rats treated with cortisone alone. However, an increase in cell-associated uPA was observed in rats with PCP. This increase appears to be produced in response to P carinii infection. In fact, when AMs obtained from untreated healthy or immunosuppressed uninfected rats were challenged with P carinii, a significant increase in PA activity in cell lysates was observed, though a lower response was obtained in cortisone-treated animals. Our results suggest that healthy AMs respond to the presence of P carinii with an increase in uPA production and that this response in immunodepressed rat-AMs is partially impaired.     

New insights into DHFR interactions: analysis of Pneumocystis carinii and mouse DHFR complexes with NADPH and two highly potent 5-(omega-carboxy(alkyloxy) trimethoprim derivatives reveals conformational correlations with activity and novel parallel ring stacking interactions

Structural data are reported for two highly potent antifolates, 2,4-diamino-5-[3',4'-dimethoxy-5'-(5-carboxy-1-pentynyl)]benzylpyrimidine (PY1011), with 5000-fold selectivity for Pneumocystis carinii dihydrofolate reductase (pcDHFR), relative to rat liver DHFR, and 2,4-diamino-5-[2-methoxy-5-(4-carboxybutyloxy)benzyl]pyrimidine (PY957), that has 80-fold selectivity for pcDHFR. Crystal structures are reported for NADPH ternary complexes with PY957 and pcDHFR, refined to 2.2 A resolution; with PY1011 and pcDHFR, refined to 2.0 A resolution; and with PY1011 and mouse DHFR (mDHFR), refined to 2.2 A resolution. These results reveal that the carboxylate of the omega-carboxyalkyloxy side chain of these inhibitors form ionic interactions with the conserved Arg in the substrate binding pocket of DHFR. These data suggest that the enhanced inhibitory activity of PY1011 compared with PY957 is, in part, due to the favorable contacts with Phe69 of pcDHFR by the methylene carbons of the inhibitor side chain that are oriented by the triple bond of the 1-pentynyl side chain. These contacts are not present in the PY957 pcDHFR complex, or in the PY1011 mDHFR complex. In the structure of mDHFR the site of Phe69 in pcDHFR is occupied by Asn64. These data also revealed a preference for an unusual parallel ring stacking interaction between Tyr35 of the active site helix and Phe199 of the C-terminal beta sheet in pcDHFR and by Tyr33 and Phe179 in mDHFR that is independent of bound ligand. A unique His174-His187 parallel ring stacking interaction was also observed only in the structure of pcDHFR. These ring stacking interactions are rarely found in any other protein families and may serve to enhance protein stability.     

Pneumocystis Carinii Pneumonia: Relationship Between Cyst Size and Response to Treatment

Pneumocystis carinii pneumonia is a common cause of death in patients with AIDS. Diagnosis is based on cytological examination of smears prepared from induced sputum samples and bronchoalveolar lavage (BAL) specimens which have been stained with methenamine silver. We have examined 46 BAL/induced sputum specimens from patients who had clinical symptoms and signs suggestive of P. carinii pneumonia and measured the diameter of a minimum 100 cysts in each specimen. We found that cyst size correlated with response to treatment with co-trimoxazole. This observation has implications for the therapeutic management of patients with this infection. La pneumopathie áPneumocystis carinii est une cause de décès fréquente chez les patients atteints d'un syndrome d'immunodéficience acquise (SIDA). Le diagnostic est basé sur l'examen cytologique des frottis préparés à partir du produit d'expectoration provoquée ou du liquide de lavage bronchoalvéolaire (LBA), après coloration de Grocott. Le diamètre d'au moins 100 kystes par échantillon a été mesuré dans 46 prélèvements de LBA/expectoration provoquée de patients présentant des signes cliniques suggérant une pneumopathie àPneumocystis carinii. Nous avons constaté que le diamètre des kystes est corrélé avec la réponse au traitment par co-trimoxazole, Cette observation a des implications sur la conduite du traitement des patients porteurs de cette infection. PCP ist eine häufige Todesursache bei HIV-Patienten. Die diagnose beruht auf dem cytologischen Nachweis in induzierten Sputumausstrichen oder bronchio-lo-alveolären Lavagen mit der Grokott-Färbung. In 46 positiven Fällen wurde der Durchmesser von mindestens 100 Cysten in jeder Probe gemessen und eine Korrelation zum Behandlungserfolg mit Co-Trimoxazol nachgewiesen.     

Unique Telomeric Expression Site of Major-Surface-Glycoprotein Genes of Pneumocystis carinii

Major cell surface glycoproteins (MSG) of Pneumocystis cariniiplay a crucial role in the host-parasite interactions involvedin P. carinii pneumonia in AIDS patients. Genes encoding MSGsare repeated, highly polymorphic, and distributed among allof the 14-15 chromosomes. Here we show, by BAL-31 exonucleasecleavage and DNA cloning experiments, that the unique expressionsite (previously termed UCS) of MSG genes located in the 500-kbchromosome is telomeric. The 11-kb genomic UCS fragment isolatedand sequenced in this study contained one MSG coding sequence(termed msg105), subtelomeric repetitive sequences and telomere-specifictandem repeats of TTAGGG oriented 5' to 3' towards the DNA end.Despite the N-terminal polymorphism, the C-terminal one-thirdsequence of MSG105 was identical to one of the known MSG-cDNAs,suggesting homologous recombination within the MSG coding sequences.These features closely resemble the Variant Surface Glycoproteinsystem of the protozoan parasite Trypanosoma brucei, suggestingthat the genetic heterogeneity of MSGs is generated by recombinationbetween the UCS expression site and multiple MSG genes by meansof reciprocal exchange or gene conversion.