Clinical aspects of pelvic inflammatory disease

Pelvic inflammatory disease (PID) is a common and poorly managed condition. Untreated or inadequately treated, it leads to tubal infertility, ectopic pregnancy and chronic pelvic pain. Diagnostic difficulties are compounded by the wide variety of clinical presentations and the insensitivity and poor specificity of laboratory tests. Better recognition of mild and atypical disease needs a high index of suspicion whenever young, sexually active women present with gynaecological symptoms. Laparoscopy supplemented by microbiological tests and fimbrial minibiopsy should be regarded as the diagnostic 'gold standard' for research studies; new studies are required to identify techniques which might reduce under- and over-diagnosis. Early treatment reduces the risk of an adverse effect on fertility. Any therapeutic regimen selected should be effective against the common aetiological agents Chlamydia trachomatis, Neisseria gonorrhoeae, genital mycoplasmas and aerobic and anaerobic bacteria. Since at least 60% of cases of PID can be attributed to infection with a sexually transmitted organism, partner notification forms an essential part of management.      

Porcine colostrum and milk stimulate visceral organ and skeletal muscle protein synthesis in neonatal piglets.

Our objective was to determine the relative contributions of protein synthesis and protein absorption in the rapid accretion of gastrointestinal protein in suckling piglets during the early neonatal period. We measured the rates of tissue protein synthesis using a flooding dose of L-[4-3H]phenylalanine in various visceral and peripheral tissues of neonatal piglets fed water, mature milk or colostrum for 6 h. The jejunal and ileal protein synthesis rates in piglets fed either colostrum or milk were three- to fourfold higher than in piglets fed water. The increased jejunal and ileal protein synthesis could not, however, account for the differences in protein mass between the colostrum-fed and water-fed groups. The relative abundance of IgG, a major porcine colostral protein, in jejunal tissue was markedly higher in piglets fed colostrum than in piglets fed either milk or water. The fractional protein synthesis rates in liver, kidney, spleen and skeletal muscle and the absolute protein synthesis rates in liver and spleen were also greater in piglets fed colostrum than in those fed milk or water. Increased endogenous protein synthesis made only a minor contribution to the increased intestinal protein accretion in neonatal piglets fed colostrum. A much larger proportion of this increase seemed to be a result of absorption and retention of ingested immunoglobulins.     

Detection of human papillomavirus in the prostate by polymerase chain reaction and in situ hybridization.

Human papillomavirus is associated with a variety of anogenital lesions, including genital warts, precancers and cancers. In male patients human papillomavirus has been identified in proliferative lesions ranging from penile and urethral warts to penile and prostatic cancers. We examined the association of human papillomavirus deoxyribonucleic acid (DNA) in 84 prostate tissue specimens. Specimens were selected from radical prostatectomy, transurethral resection or transrectal biopsy procedures. A total of 60 formalin-fixed, paraffin-embedded tissues (24 prostate cancer specimens, 16 benign prostatic hyperplasia specimens and 20 normal specimens) was examined by polymerase chain reaction and in situ hybridization. Also, 24 gelatin-embedded frozen prostate cancer specimens were examined for human papillomavirus DNA by polymerase chain reaction. Of the specimens 69 were deemed adequate for polymerase chain reaction analysis, whereas all 60 paraffin-embedded tissues were sufficient for in situ hybridization. Human papillomavirus DNA was detected in 2 normal tissues and 6 prostate cancers using polymerase chain reaction. None of the benign prostatic hyperplasia specimens was positive for human papillomavirus. Human papillomavirus typing results indicated that virus type 16 was present in each of the 8 positive specimens. Confirmation of the presence of human papillomavirus was obtained for 1 of the prostate cancers by nonisotopic in situ hybridization with biotinylated human papillomavirus genomic probes. The low prevalence of human papillomavirus in this study population does not strongly support an etiological role for the virus in prostate cancer.     

Incidence and determinants of moderate COPD (GOLD II) in male smokers aged 40–65 years: 5-year follow up

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a major health problem with an estimated prevalence of 10-15% among smokers. The incidence of moderate COPD, as defined by the Global Initiative for Chronic Obstructive Lung Disease (GOLD), is largely unknown. AIM: To determine the cumulative incidence of moderate COPD (forced expiratory volume in 1 second/forced vital capacity ratio [FEV1/FVC] <0.7 and FEV1 <80% predicted) and its association with patient characteristics in a cohort of male smokers. DESIGN: Prospective cohort study. SETTING: The city of IJsselstein, a small town in the Netherlands. METHOD: Smokers aged 40-65 years who were registered with local GPs, participated in a study to identify undetected COPD. Baseline measurements were taken in 1998 of 399 smokers with normal spirometry (n = 292) or mild COPD (FEV1/FVC <0.7 and FEV1 >or=80% predicted, n = 107) and follow-up measurements were conducted in 2003. RESULTS: After a mean follow-up of 5.2 years, 33 participants developed moderate COPD (GOLD II). This showed an estimated cumulative incidence of 8.3% (95% CI = 5.8 to 11.4) and a mean annual incidence of 1.6%. No participant developed severe airflow obstruction. The risk of developing moderate COPD in smokers with baseline mild COPD (GOLD I) was five times higher than in those with baseline normal spirometry (one in five versus one in 25). CONCLUSIONS: In a cohort of middle-aged male smokers, the estimated cumulative incidence of moderate COPD (GOLD II) over 5 years was relatively high (8.3%). Age, childhood smoking, cough, and one or more GP contacts for lower respiratory tract problems were independently associated with incident moderate COPD.     

Chest radiography in general practice: indications, diagnostic yield and consequences for patient management

BACKGROUND: Chest radiography (CXR) is frequently performed in Western societies. There is insufficient knowledge of its diagnostic value in terms of changes in patient management decisions in primary care. AIM: To assess the influence of CXR on patient management in general practice. DESIGN OF STUDY: Prospective cohort study. SETTING: Seventy-eight GPs and three general hospitals in the Netherlands. METHOD: Patients (n = 792) aged > or =18 years referred by their GPs for CXR were included. The main outcome was change in patient management assessed by means of questionnaires filled in by GPs before and after CXR. RESULTS: Mean age of the patients was 57.3+/-16.2 years and 53% were male. Clinically relevant abnormalities were found in 24% of the CXRs. Patient management changed in 60% of the patients following CXR. Main changes included: fewer referrals to a medical specialist (from 26 to 12%); reduction in initiation or change in therapy (from 24 to 15%); and more frequent reassurance (from 25 to 46%). However, this reassurance was not perceived as such in a quarter of these patients. A change in patient management occurred significantly more frequently in patients with complaints of cough (67%), those who exhibited abnormalities during physical examination (69%), or those with a suspected diagnosis of pneumonia (68%). CONCLUSION: Patient management by the GP changed in 60% of patients following CXR. CXR substantially reduced the number of referrals and initiation or change in therapy, and more patients were reassured by their GP. Thus, CXR is an important diagnostic tool for GPs and seems a cost-effective diagnostic test.     

Analysis of the effect of DNA purification on detection of human papillomavirus in oral rinse samples by PCR

Human papillomavirus (HPV) has recently been associated with oral cancers. To prepare for a study of the natural history of oral HPV infection, the effect of the DNA purification method on HPV genomic DNA detection in Scope mouthwash oral rinse samples and the reproducibility of HPV detection in rinse samples collected 7 days apart were investigated. The study was conducted with a population at high risk for oral HPV infection: human immunodeficiency virus-infected men with CD4-cell counts <200. Five DNA purification methods were compared among equal aliquots of oral rinse samples collected from a subset of individuals. The purification methods included (i) proteinase K digestion (PKD) and heat inactivation; (ii) PKD and ethanol precipitation (EP); (iii) PKD, phenol-chloroform extraction, and EP; (iv) use of the Puregene DNA purification kit; and (v) use of the QIAamp DNA Blood Midi kit. HPV was detected by PCR amplification with PGMY09 and PGMY11 L1 primer pools and by use of a Roche linear array. Puregene-purified samples had higher human DNA yields and purities, and Puregene purification detected the greatest number of HPV-positive subjects and total HPV infections in comparison to the numbers detected by all other methods. The total number of HPV infections and HPV prevalence estimates were also higher for Puregene-processed oral rinse samples when a fixed volume (10 mul) rather than a fixed cell number ( approximately 50,000 cells) was used for PCR amplification. A good concordance was observed for oral HPV infection status (agreement, 80%; kappa value, = 0.60) and type-specific infection (agreement, 98%; kappa value, 0.57) in matched oral rinse samples. The method of DNA purification significantly affects the detection of HPV genomic DNA from oral rinse samples and may result in exposure misclassification that could contribute to the inconsistent associations reported in the literature.     

Improved amplification of genital human papillomaviruses

Genital human papillomaviruses (HPVs) are commonly detected from clinical samples by consensus PCR methods. Two commonly used primer systems, the MY09-MY11 (MY09/11) primers and the GP5+-GP6+ (GP5+/6+) primers, amplify a broad spectrum of HPV genotypes, but with various levels of sensitivity among the HPV types. Analysis of the primer-target sequence homology for the MY09/11 primers showed an association between inefficient amplification of HPV types and the number and position of mismatches, despite accommodation of sequence variation by inclusion of degenerate base sites. The MY09/11 primers were redesigned to increase the sensitivity of amplification across the type spectrum by using the same primer binding regions in the L1 open reading frame. Sequence heterogeneity was accommodated by designing multiple primer sequences that were combined into an upstream pool of 5 oligonucleotides (PGMY11) and a downstream pool of 13 oligonucleotides (PGMY09), thereby avoiding use of degenerate bases that yield irreproducible primer syntheses. The performance of the PGMY09-PGMY11 (PGMY09/11) primer system relative to that of the standard MY09/11 system was evaluated with a set of 262 cervicovaginal lavage specimens. There was a 91.5% overall agreement between the two systems (kappa = 0.83; P < 0.001). The PGMY09/11 system appeared to be significantly more sensitive than the MY09/11 system, detecting an additional 20 HPV-positive specimens, for a prevalence of 62.8% versus a prevalence of 55.1% with the MY09/11 system (McNemar's chi(2) = 17.2; P < 0.001). The proportion of multiple infections detected increased with the PGMY09/11 system (40. 0 versus 33.8% of positive infections). HPV types 26, 35, 42, 45, 52, 54, 55, 59, 66, 73, and MM7 were detected at least 25% more often with the PGMY09/11 system. The PGMY09/11 primer system affords an increase in type-specific amplification sensitivity over that of the standard MY09/11 primer system. This new primer system will be useful in assessing the natural history of HPV infections, particularly when the analysis requires HPV typing.     

Potent induction of human colon cancer cell uptake of chemotherapeutic drugs by N-myristoylated protein kinase C-α (PKC-α) pseudosubstrate peptides through a P-glycoprotein-independent mechanism

Phorbol ester protein kinase C (PKC) activators and PKC isozyme over-expression have been shown to significantly reduce intracellular accumulation of chemotherapeutic drugs, in association with the induction of multidrug resistance (MDR) in drug-sensitive cancer cells and enhancement of drug resistance in MDR cancer cells. These observations constitute solid evidence that PKC plays a significant role in the MDR phenotype of cancer cells. PKC-catalyzed phosphorylation of the drug-efflux pump P-glycoprotein was recently ruled out as a contributing factor in MDR. At present, the sole drug transport-related event that has been identified as a component of the role of PKC in MDR is PKC-induced expression of the P-glycoprotein-encoding gene mdr1. The objective of this study was to test the hypothesis that PKC can modulate the uptake of chemotherapeutic drugs in cancer cells independently of P-glycoprotein. We analyzed the effects of selective PKC activators/inhibitors on the uptake of radiolabelled cytotoxic drugs by cultured human colon cancer cells that lacked P-glycoprotein activity and did not express the drug efflux pump at the level of message (mdr1) or protein. We found that the selective PKC activator 12-O-tetradecanoylphorbol-13-acetate (TPA) significantly reduced uptake of [¹⁴C] Adriamycin and [³H] vincristine in human colon cancer cells devoid of P-glycoprotein activity, and that PKC-inhibitory N-myristoylated PKC- pseudosubstrate synthetic peptides potently and selectively induced uptake of the cytotoxic drugs in the phorbol ester-treated and non-treated colon cancer cells. TPA treatment of the cells did not induce expression of either P-glycoprotein or its message mdr1. In contrast with [¹⁴C]Adriamycin and [³H] vincristine uptake, [³H] 5-fluorouracil uptake by the cells was unaffected by TPA and reduced by the PKC-inhibitory peptides. These results indicate that PKC activation can significantly reduce the uptake of multiple cytotoxic drugs by cancer cells independently of P-glycoprotein, and that N-myristoylated PKC- pseudosubstrate peptides potently and selectively induce uptake of multiple cytotoxic drugs in cultured human colon cancer cells by a novel mechanism that does not involve P-glycoprotein and may involve PKC isozyme inhibition. Thus, N-myristoylated PKC- pseudosubstrate peptides may offer a basis for the development of agents that reverse intrinsic drug resistance in human colon cancer.