Impact of accelerated progression to AIDS on public health monitoring of late HIV diagnosis

Some patients develop AIDS within a year of HIV infection ("accelerated progression"). Classifying such cases as late HIV diagnosis may lead to inaccurate evaluation of HIV testing efforts. We sought to determine this group's contribution to overall late diagnosis rates. To identify cases of accelerated progression (development of AIDS within 12 months of a negative HIV test), we reviewed published HIV seroconverter cohort studies and used New York City's (NYC) HIV/AIDS surveillance registry. From the literature review, three seroconverter cohort studies revealed that 1.0-3.6% of participants had accelerated progression to AIDS. Applying this frequency estimate to the number of new infections in NYC (4762) for 2006 calculated by the Centers for Diseases Control and Prevention's incidence formula, we estimated that 3.6-13.0% of 1317 NYC HIV cases who are diagnosed with AIDS within 12 months of HIV diagnosis are accelerated progressors, not persons HIV infected for many years who did not test and present with AIDS (i.e., delayed diagnosis). In addition, our analysis of the 2006 NYC surveillance registry confirmed the occurrence of accelerated progression in a population-based setting; 67 accelerated progressors were reported and 9 (13%) could be confirmed through follow-up medical record review. With increased HIV testing initiatives, the irreducible proportion of AIDS cases with accelerated progression must be considered when interpreting late diagnosis data.     

Serum factors in older individuals change cellular clock properties

Human aging is accompanied by dramatic changes in daily sleep-wake behavior: Activity shifts to an earlier phase, and the consolidation of sleep and wake is disturbed. Although this daily circadian rhythm is brain-controlled, its mechanism is encoded by cell-autonomous circadian clocks functioning in nearly every cell of the body. In fact, human clock properties measured in peripheral cells such as fibroblasts closely mimic those measured physiologically and behaviorally in the same subjects. To understand better the molecular mechanisms by which human aging affects circadian clocks, we characterized the clock properties of fibroblasts cultivated from dermal biopsies of young and older subjects. Fibroblast period length, amplitude, and phase were identical in the two groups even though behavior was not, thereby suggesting that basic clock properties of peripheral cells do not change during aging. Interestingly, measurement of the same cells in the presence of human serum from older donors shortened period length and advanced the phase of cellular circadian rhythms compared with treatment with serum from young subjects, indicating that a circulating factor might alter human chronotype. Further experiments demonstrated that this effect is caused by a thermolabile factor present in serum of older individuals. Thus, even though the molecular machinery of peripheral circadian clocks does not change with age, some age-related circadian dysfunction observed in vivo might be of hormonal origin and therefore might be pharmacologically remediable.     

Six novel mutations in the proopiomelanocortin and melanocortin receptor 4 genes in severely obese adults living in southern Italy

BACKGROUND: The genetic characterization of obese individuals could clarify the molecular mechanisms underlying body weight regulation and lead to targeted therapy. Here we report variants of the proopiomelanocortin (POMC) and melanocortin receptor 4 (MC4R) genes detected in severely obese adults living in southern Italy. METHODS: A total of 196 unrelated nondiabetic severely obese individuals [111 females and 85 males; mean (SD) age, 32.2 (11.5) years; mean body mass index, 48.8 (8.1) kg/m(2)] and 100 normal-weight healthy volunteers (34 males and 66 females) entered the study. POMC and MC4R were genotyped by sequencing analysis. Leptin, insulin, glucose, and the lipid profile were measured in fasting serum samples. We used the protein truncation test to verify the stop-codon mutation. Anthropometric measurements, sitting blood pressure, and heart rate were also recorded. RESULTS: Of the obese participants, 1.5% had mutations in POMC exon 3 (new mutations, P231L and E244X; known, R236G) and 2.5% had MC4R mutations (new mutations, W174C, Q43X, S19fsX51, and I317V; known, A175T). These mutations were not present in the controls. Gene polymorphisms were identified in similar percentages of severely obese and nonobese individuals, i.e., respectively, 52.5% and 51% (POMC) and 1% and 2% (MC4R). CONCLUSIONS: We detected 2 new POMC mutations and 4 new MC4R mutations in a large number of severely obese adults living in southern Italy. These mutations, not present in normal-weight individuals, are further evidence that defects in the melanocortin pathway are related to severe obesity.     

High Prevalence of qnr Genes in Commensal Enterobacteria from Healthy Children in Peru and Bolivia

A remarkable prevalence of qnrB (54%) and, at a lower level, of qnrS (14%) was discovered in pools of commensal enterobacteria from 310 healthy children living in Peru and Bolivia, using a metagenomic approach. Analysis of randomly selected enterobacterial pools revealed that qnrB was mainly carried by Escherichia coli and qnrS by Klebsiella pneumoniae. Investigation of 11 qnrB-positive isolates and 9 qnrS-positive isolates revealed the presence of plasmid-borne qnrB19 (n = 8), qnrB2 (n = 2), qnrB10 (n = 1), and qnrS1 (n = 9) genes.Several plasmid-mediated quinolone resistance (PMQR) mechanisms have been discovered during the past decade, including Qnr proteins, QepA transporters, and the acetyltransferase AAC(6′)-Ib-cr (11, 13). Qnr proteins, the first discovered PMQR mechanism, are small pentapeptide repeat proteins that bind and protect type II DNA topoisomerases from inhibition by fluoroquinolones (17-19). Different lineages of Qnr proteins have been described (QnrA, QnrB, QnrS, and, more recently, QnrC and QnrD), with several allelic variants known for some of them (3, 9, 11, 20). qnr-like genes have also been detected on chromosomes from both gram-negative and gram-positive bacteria (9), and, recently, as class 1 integron gene cassettes in the chromosome of Vibrio cholerae (7). Although Qnr proteins only determine a moderate reduction of quinolone susceptibility, this may favor the selection of additional resistance mechanisms leading to higher-level quinolone resistance of clinical significance, and the dissemination of qnr genes and other PMQR determinants is believed to be an important promoter for evolution of quinolone resistance (11-13).qnr genes have been reported worldwide, especially in enterobacteria. However, they have mostly been investigated in clinical isolates with specific resistance traits (e.g., quinolone resistance or extended-spectrum β-lactamase phenotype) (11 and references therein), while their prevalence in commensal bacteria remains largely unknown. In this study, we investigated the prevalence of qnr genes in commensal enterobacteria from healthy children by a PCR-based metagenomic approach. We also tested a simplified dot blot DNA hybridization method as a less-labor-intensive and expensive tool to perform the metagenomic analysis.The analysis was carried out on commensal enterobacterial pools from 310 healthy children, ages 6 to 72 months, living in four urban areas of Latin America: two in Peru (Moyobamba, San Martin Department; and Yurimaguas, Loreto Department) and two in Bolivia (Camiri, Santa Cruz Department; and Villa Montes, Tarija Department). The enterobacterial pools, consisting of the bacterial growth obtained by plating fecal samples (one sample per child) onto MacConkey agar (MCA) plates (Oxoid, Milan, Italy), were randomly selected among samples (n = 3,193) obtained during a survey performed in 2005 in the same areas (1) and stored at −70°C. A loopful of each pool was plated on an MCA plate supplemented with 0.12 μg/ml ciprofloxacin (MCA-CIP). This ciprofloxacin concentration was used for screening purposes since (i) it was lower than MICs usually exhibited by enterobacterial strains harboring qnr genes as the sole quinolone resistance mechanism (reference 11 and references therein), while being higher than the wild-type MIC distribution for Escherichia coli and Klebsiella pneumoniae (http://www.escmid.org/research_projects/eucast/); (ii) a similar ciprofloxacin MIC threshold has previously been used for screening of qnr-positive bacteria (4, 8, 14, 15, 21). In case of growth onto MCA-CIP, a loopful of bacteria was directly used for total DNA extraction (10), and about 100 ng of metagenomic DNA was used as template in PCRs (50 μl) to detect the presence of qnr genes. Primers and conditions for PCR amplification of qnr genes were described previously in references 14 (qnrA and qnrS) and 2 (qnrB). Controls for qnr genes were kindly provided by Patrice Nordmann and Laurent Poirel (Université Paris-Sud, K.-Bicêtre, France). The specificity of the PCR products was confirmed by partial sequence analysis of randomly selected amplicons (Macrogen, Seoul, Korea).The presence of qnr genes in the enterobacterial pools was also investigated by dot blot DNA hybridization using a rapid method, essentially as described by Srinivasan et al. (16). Briefly, a loopful of the bacterial growth on MCA-CIP was transferred to 150 μl of lysis solution (0.4 N NaOH, 10 mM EDTA) and incubated at 70°C for 2 h. The bacterial lysate (100 μl) was directly blotted onto Hybond-N+ nylon membranes (Amersham Bioscience, Buckinghamshire, United Kingdom) using a BIO-dot microfiltration apparatus (Bio-Rad Laboratories, Milan, Italy). Nylon membranes were washed twice with 2× SSC (1× SSC is 0.15 M NaCl plus 0.015 M sodium citrate), fixed by UV light, and hybridized with digoxigenin-dUTP (DIG)-labeled qnr probes (generated by PCR using the positive controls, as described above) using the DIG system according to the manufacturer''s instructions (Roche Diagnostics SpA, Milan, Italy).Growth of enterobacterial pools on MCA-CIP was observed with 275 of 310 samples (89%). Analysis of the metagenomes prepared from this bacterial growth by PCR revealed overall positivities of 54% for qnrB and 14% for qnrS, while qnrA was not detected (Table ​(Table1).1). Dot blotting showed signals of variable intensity (Fig. ​(Fig.1)1) and an overall lower detection sensitivity, but results were fully consistent with those of PCR (i.e., all positive samples in the dot blot, including weak signals, were also positive by PCR) (Table ​(Table1).1). This evidently reflected a variable copy number of target genes in the metagenomic DNA from various samples, with a number of cases in which the amount of target genes could only be detected by the more sensitive gene amplification approach.Open in a separate windowFIG. 1.Nylon membrane prepared with bacterial lysates, hybridized with DIG-labeled qnrB probe. All positive signals, including the weakest ones, were positive in PCR experiments. Positive and negative controls (C+ and C−, respectively) are indicated by arrows.

TABLE 1.

Prevalence of qnr genes in commensal enterobacteria from 310 healthy children living in Peru and Bolivia^a

Earliest Videofluoromanometric Pharyngeal Signs of Dysphagia in ALS Patients

The aim of this study was to find whether there are manometric pharyngeal changes that may have diagnostic and prognostic relevance in the amyotrophic lateral sclerosis (ALS) patient who does not show changes in contrast-medium oropharyngeal transit in a videofluoroscopic swallowing study. Ten ALS patients, with an ALS Severity Scale Score of at least 7, no need to change dietary habit, no aspiration and/or penetration, and no other changes in contrast-medium oropharyngeal transit, were collected from our institution’s database of videofluoromanometric swallowing studies. They were included in the study together with a group of 11 healthy volunteers. For each subject, 12 manometric items—7 for the pharyngeal phase and 5 for UES functionality—were evaluated. Statistically significant differences between the ALS patients and the healthy volunteers were found for pharyngeal contraction time of the upper region (median = 1,120, range = 880–1,420 vs. median = 970, range = 800–1,140), pharyngeal contraction time of the intermediate region (median = 1140, range = 960–1,360 vs. median = 770, range = 280–1,180), pharyngeal contraction time of the lower region (median = 1,320, range = 920–1,760 vs. median = 800, range = 620–1,780), and residual pressure after the relaxation of the UES (median = 2.2, range = ?20.2 to 27.8 vs. median = ?5.7, range = ?2.9 to 8.4). A videofluoromanometric swallowing study may show an increase in the pharyngeal contraction time and in residual pressure after relaxation of the UES in ALS patients without videofluoroscopic changes in contrast-medium oropharyngeal transit.     

Immunohistochemical detection of p21ras, c-myc and p53 oncoproteins in hepatocellular carcinoma and in non-neoplastic liver tissue

BACKGROUND: Genetic and epigenetic alterations have been described in animal hepatocarcinogenesis models but need to be studied in human being. AIMS: To assess the immunoreactivity of p21ras, c-myc and p53 oncoproteins in hepatocellular carcinoma and non neoplastic tissue. Association of the immunoreactivity of these markers with histological grades and patterns, hepatitis B and C were additionally studied. METHODS: Detection of oncoproteins p21ras, c-myc and p53 was performed immunohistochemically in hepatocellular carcinoma (47 cases) and surrounding non neoplastic liver tissue (40 cases). RESULTS: Oncoproteins p21ras, c-myc and p53 were detected in 44,7%, 53,2% and 36,2% of the hepatocellular carcinoma cases, respectively. The p21ras and c-myc immunoreactivity has shown a significant association. However there was no association of p21ras, c-myc and p53 detection with hepatitis B and C virus infections, histological grades and patterns. The same significant association between p21ras and c-myc was observed in non-neoplastic tissue with cirrhosis when compared with tissue without it. The p53 immunoreactivity was negative in all non-neoplastic liver tissue samples. CONCLUSIONS: The immunoreactivity detection of p21ras, c-myc and p53 corroborates previous evidence of their detection in hepatocellular carcinoma that suggest the participation of these proteins in human hepatocarcinogenesis. The significant association between p21ras and c-myc oncoproteins in hepatocellular carcinoma and in cirrhosis can point to an interaction between them mainly, in hepatocarcinogenesis that occurs through cirrhosis.     

Prevalence,characteristics and outcomes of older patients with hereditary versus wild-type transthyretin amyloid cardiomyopathy

Aims

Transthyretin amyloid cardiomyopathy (ATTR-CM) is often assumed to be associated with wild-type TTR genotype (ATTRwt) in elderly patients (aged ≥70), some of whom are not offered genetic testing. We sought to estimate the prevalence, clinical characteristics and prognostic implications of transthyretin (TTR) variants among elderly patients diagnosed with ATTR-CM.

Methods and results

Data from consecutive patients over 70 years of age diagnosed with ATTR-CM at the UK National Amyloidosis Centre between January 2010 and August 2022 were retrospectively evaluated. All patients underwent clinical evaluation, biochemical tests, echocardiography and TTR genotyping. The study outcome was all-cause mortality. The study population consisted of 2029 patients with ATTR-CM (median age 79 years at diagnosis, 13.5% females, 80.4% Caucasian). Variant ATTR-CM (ATTRv-CM) was diagnosed in 20.7% (n = 421) of the study population of whom 327 (77.7%) carried V122I, 47 (11.2%) T60A, 16 (3.8%) V30M and 31 (7.3%) other pathogenic TTR variants. During a median (range) follow-up of 29 (12–48) months, ATTRv-CM was associated with increased all-cause mortality compared to ATTRwt-CM, with the poorest survival observed in V122I-associated ATTRv-CM (p < 0.001). Univariable and multivariable logistic regression analyses in those with ATTR-CM showed younger age at diagnosis (odds ratio [OR] 0.85 per year, p < 0.001), female sex (OR 2.73, p < 0.001), Afro-Caribbean ethnicity (OR 65.5, p < 0.001), atrial fibrillation (OR 0.65, p = 0.015), ischaemic heart disease (OR 0.54, p = 0.007), peripheral polyneuropathy (OR 5.70, p < 0.001) and orthostatic hypotension (OR 6.29, p < 0.001) to be independently associated with ATTRv-CM.

Conclusion

Up to 20.7% of elderly patients with ATTR-CM have a pathogenic TTR variant. These findings support routine sequencing of the TTR gene in all patients with ATTR-CM regardless of age.     

Variants of CARD15 are associated with an aggressive clinical course of Crohn's disease--an IG-IBD study

BACKGROUND: Three major variants of the CARD15 gene confer susceptibility to Crohn's disease (CD). Whether or not these variants correlate with specific clinical features of the disease is under evaluation. AIM: We investigated the possible association of CARD15 variants with specific clinical characteristics, including the occurrence of anti-Saccharomyces cerevisiae antibodies (ASCA) and antineutrophil cytoplasmic antibodies (ANCA), in a large cohort of inflammatory bowel disease (IBD) patients and their unaffected relatives. METHODS: Three hundred and sixteen CD patients (156 with positive family history), 408 ulcerative colitis (UC) patients (206 with positive family history), 588 unaffected relatives, and 205 unrelated healthy controls (HC) were studied. Single nucleotide polymorphisms (SNPs) R702W, G908R, and L1007finsC of the CARD15 gene were investigated and correlated to age at diagnosis, gender, family history, localization, extraintestinal manifestations, previous resective surgery, stenosing/fistulizing pattern, ANCA, and ASCA. RESULTS: Compared to HC, the frequencies of all three variants in CD were significantly increased: 8.7% versus 4.1% for R702W (p < 0.006), 7.3% versus 2.7% for G908R (p < 0.002), 9.3% versus 0.7% for L1007finsC (p < 0.00001). At least one risk allele was found in 38.2% (p < 0.0001, compared to HC), 13.7% (NS), and 15.1% of CD, UC, and HC, respectively. The L1007finsC risk allele was also significantly increased in unaffected relatives of familial (9.5%; p < 0.00001), and sporadic CD (9%; p < 0.00001), compared to HC (0.7%). Sixteen healthy relatives, carriers of two risk alleles, were asymptomatic after 5-8 yr of follow-up. CD carriers of at least one variant were younger (p= 0.03), more likely to have ileal localization (p= 0.0001), stenosing pattern (p= 0.01), previous resective surgery (p= 0.0001), and presence of ASCA (p= 0.0001). No difference in SNPs frequency between familial and sporadic cases of CD was found. CONCLUSION: In our population, both familial and sporadic CD patients carrying at least one major variant of CARD15 had an aggressive clinical course.     

Clinical significance of the quantitative assessment of the cytosolic concentration of HER-2/neu protein in breast cancer by immunoenzymatic assay (ELISA)

Purpose: Retrospective analysis to assess the prognostic and predictive value of HER-2/ neu expression in breast tumors, quantified by enzyme immunoassay (ELISA).Methods: Quantification of HER-2/neu was performed on cytosolic extracts from 914 cases of primary invasive breast carcinomas. Relapse-free and overall survival data were available from 889 patients. The prognostic value of HER-2/neu levels was assessed considering them as a continuous, dichotomic or quartile variable.Results: Cytosolic HER-2/neu levels ranged widely in breast carcinomas (median: 746.5 NHU/mg; range: 2.8–80,000 NHU/mg protein). HER-2/neu protein levels were significantly higher in either moderately or poorly differentiated tumors, as well as in those showing a ductal histological type, aneuploidy or a high S-phase fraction. There was a significant and positive association between cytosolic and membranous HER-2/neu levels (n=162, r sub S=0.53; P<0.0001). In addition, cytosolic HER-2/neu level correlated weakly with progesterone receptors but not with estrogen receptors. Elevated cytosolic HER-2/neu levels (≥1,400 NHU/mg protein) were associated with a high probability of both shortened relapse-free survival and overall survival. This same cut-off value was obtained when we divided the overall group of patients in a training set. However, this HER-2/neu value did not achieve any statistical significance in a validation set used to make sure that the cut-off was correct. Nevertheless, when we divided the obtained data into three different groups with respect to the quartile values (Q) of the intratumoral oncoprotein levels (≤ Q ₁ vs Q ₁−Q ₂ vs > Q ₃), we observed that patients with either low HER-2/ neu levels (≤ Q ₁) or high HER-2/neu levels (> Q ₃) had shorter both relapse-free survival and overall survival curves than those patients with intermediate HER-2/neu levels. On the other hand, high HER-2/neu levels predicted a poor response to adjuvant chemotherapy but not to adjuvant hormonal therapy with tamoxifen.Conclusions: The results of the present investigation indicate that by quantitatively determining the content of HER-2/neu oncoprotein, groups of high-risk breast cancer patients could be identified, for a more effective clinical management.     

Treatments of AIDS-related Kaposi's sarcoma

Although Kaposi's sarcoma (KS) has decreased in countries where the highly active antiretroviral therapy (HAART) regimen is available, however it remains, after non-Hodgkin's lymphomas, the most common malignancy in HIV+ patients. Advances in the treatment of AIDS-KS have been achieved, even though a gold standard therapy has not been yet defined. With the availability of HAART, a dramatic KS clinical response has been documented, making HAART essential in all patients. In case of aggressive and/or life threatening KS, more complex therapeutic schedules have to be taken into account, including chemotherapy and/or immunotherapy. Liposomal anthracyclines and paclitaxel have been approved by FDA as first line and second line mono-therapy, respectively. Interferon-alpha (INF-alpha) is the only immunomodulant agent to have shown a therapeutic effect. Among the new drugs, many antiangiogenetic agents have produced encouraging responses. Finally, the identification of the HHV-8 as a causative agent and new metalloproteinase inhibitors may offer promising targets for the KS treatment.