Monoclonal antibody analysis of T-lymphocyte subsets in young and aged adults

Eleven monoclonal antibodies identifying surface antigens present on human T lymphocytes were utilized to investigate the effects of advanced age on peripheral blood lymphocyte subsets. Both the proportion and number of lymphocytes recognized by five antibodies reactive with 'pan' T cell antigens (OKT3, OKT11, Leu4, T101 and Lyt3) decreased with age. The percentage of helper/inducer (OKT4+, Leu3a+) cells remained constant; however the proportion of Leu2a+, suppressor/cytotoxic cells declined. There was no change with age in the percent representation of OKT9+ or OKT10+ cells, nor in the ratio of helper/inducer to suppressor/cytotoxic cells (OKT4+/OKT8+ or Leu3a+/Leu2a+). Absolute numbers of helper/inducer (OKT4+, Leu3a+), suppressor/cytotoxic (OKT8+, Leu2a+), OKT9+ and OKT10+ cells were lower in elderly individuals as the result of lymphocytes constituting a lower percentage of the peripheral blood white cell population in this age group. While only small differences existed between the lymphocyte populations of young and aged men; aged women, compared to young women, had more dramatic shifts in their T cell populations. Comparison of antibodies putatively identifying similar (the same) functional groups of T cells demonstrated excellent correlation between the percentage of cells enumerated with the antibodies OKT3+: Leu4+ (r = .951), OKT4+: Leu3a+ (r = .914), OKT8+: Leu2a+ (r = .896), and in the ratio of helper/inducer to cytotoxic/suppressor OKT4+/OKT8+: Leu3a+/Leu2a+ (r = .926) cells.     

Immunoglobulin peptide chain synthesis in the newborn rabbit

Lymphoid tissue from newborn rabbits was examined using fluorochrome conjugated goat antisera to rabbit γ, μ and α heavy chain and mixed light chain. Unimmunized newborn rabbits examined at intervals up to 20 days of age revealed a very low background of approximately one in 10⁵ cells which stained with either the anti-μ, anti-γ or anti-L chain reagents. Newborn rabbits, immunized intraperitoneally on the day of birth with S. paratyphi B (4, 5, 12:b), showed a 1000-fold increase in the number of stained cells; μ heavy chain containing cells were found 16 hours after immunization, and γ heavy chain containing cells were found 20 hours after immunization in the rabbit spleen tissue. The numbers of μ containing cells and γ containing cells were approximately equal at 5 days of age in the immunized groups. No α heavy chain containing cells were found in any of the rabbits studied. All cells which stained with the anti-μ or γ heavy chain reagent were also stained with the anti-light chain reagent.The cellular response of the immunized newborn rabbits could be inhibited by passive antibody to S. paratyphi B, either given at the time of immunization, or passively acquired from the doe. Specificity of inhibition was indicated by the failure of sheep red blood cells (SRBC) stroma to be inhibited in animals so suppressed.The range of cellular morphology of the γ chain containing and the μ chain containing cells was indistinguishable. The earliest cell to be stained with either anti-γ or anti-μ was a large blast-like cell with a thin rim of cytoplasm and a large nucleus. A full range of forms between this and typical plasma cells appeared later in each case.These findings present a paradox since γG antibody to this antigen does not appear in the serum of such animals following γM class in the usual sequence characteristic of adult animals. Possible explanations of this paradox are explored.     

Age differences in phagocytosis by polymorphonuclear leukocytes measured by flow cytometry

Elderly persons have increased morbidity and mortality due to bacterial infections. Since the polymorphonuclear leukocyte (PMN) is a major defense against bacterial infection, we utilized fluorescent microspheres and flow cytometry to examine phagocytosis by PMNs from 55 young and middle-aged adults (mean age 41.5 yrs) and two groups of elderly subjects: one group of 35 healthy individuals (mean age 74.1 years) living at home, and a second group of 11 residents (mean age 83.1 years) with severe mental and physical disabilities, living in a domiciliary care facility. We determined the percent phagocytic PMNs, the number of microspheres per PMN, and the number of microspheres per 100 PMNs. The mean number of microspheres per phagocytic PMN was similar for all groups. Statistically significant differences were found between the young and middle-aged group and the healthy or ill elderly groups for the percent phagocytic PMNs (75.3% vs 51.5% and 43.8%), and the number of microspheres per 100 phagocytic PMNs (197.3 vs 131.4 and 103.2). There were no significant differences in these parameters between healthy and debilitated elderly subjects. These data document that there is an age-related increase in representation of a population of PMNs which have a defect in phagocytic ability.     

Global analysis of outer membrane proteins from Leptospira interrogans serovar Lai

Recombinant leptospiral outer membrane proteins (OMPs) can elicit immunity to leptospirosis in a hamster infection model. Previously characterized OMPs appear highly conserved, and thus their potential to stimulate heterologous immunity is of critical importance. In this study we undertook a global analysis of leptospiral OMPs, which were obtained by Triton X-114 extraction and phase partitioning. Outer membrane fractions were isolated from Leptospira interrogans serovar Lai grown at 20, 30, and 37 degrees C with or without 10% fetal calf serum and, finally, in iron-depleted medium. The OMPs were separated by two-dimensional gel electrophoresis. Gel patterns from each of the five conditions were compared via image analysis, and 37 gel-purified proteins were tryptically digested and characterized by mass spectrometry (MS). Matrix-assisted laser desorption ionization-time-of-flight MS was used to rapidly identify leptospiral OMPs present in sequence databases. Proteins identified by this approach included the outer membrane lipoproteins LipL32, LipL36, LipL41, and LipL48. No known proteins from any cellular location other than the outer membrane were identified. Tandem electrospray MS was used to obtain peptide sequence information from eight novel proteins designated pL18, pL21, pL22, pL24, pL45, pL47/49, pL50, and pL55. The expression of LipL36 and pL50 was not apparent at temperatures above 30 degrees C or under iron-depleted conditions. The expression of pL24 was also downregulated after iron depletion. The leptospiral major OMP LipL32 was observed to undergo substantial cleavage under all conditions except iron depletion. Additionally, significant downregulation of these mass forms was observed under iron limitation at 30 degrees C, but not at 30 degrees C alone, suggesting that LipL32 processing is dependent on iron-regulated extracellular proteases. However, separate cleavage products responded differently to changes in growth temperature and medium constituents, indicating that more than one process may be involved in LipL32 processing. Furthermore, under iron-depleted conditions there was no concomitant increase in the levels of the intact form of LipL32. The temperature- and iron-regulated expression of LipL36 and the iron-dependent cleavage of LipL32 were confirmed by immunoblotting with specific antisera. Global analysis of the cellular location and expression of leptospiral proteins will be useful in the annotation of genomic sequence data and in providing insight into the biology of Leptospira.     

Cryolite glass prosthetic eyes—the response of the anophthalmic socket

Graefe's Archive for Clinical and Experimental Ophthalmology - To investigate mucoid discharge and the inflammatory response of anophthalmic sockets to cryolite glass prosthetic eye wear. A...     

Which Clinical and Patient Factors Influence the National Economic Burden of Hospital Readmissions After Total Joint Arthroplasty?

Background

The Affordable Care Act of 2010 advanced the economic model of bundled payments for total joint arthroplasty (TJA), in which hospitals will be financially responsible for readmissions, typically at 90 days after surgery. However, little is known about the financial burden of readmissions and what patient, clinical, and hospital factors drive readmission costs.

Questions/purposes

(1) What is the incidence, payer mix, and demographics of THA and TKA readmissions in the United States? (2) What patient, clinical, and hospital factors are associated with the cost of 30- and 90-day readmissions after primary THA and TKA? (3) Are there any differences in the economic burden of THA and TKA readmissions between payers? (4) What types of THA and TKA readmissions are most costly to the US hospital system?

Methods

The recently developed Nationwide Readmissions Database from the Healthcare Cost and Utilization Project (2006 hospitals from 21 states) was used to identify 719,394 primary TJAs and 62,493 90-day readmissions in the first 9 months of 2013 based on International Classification of Diseases, 9th Revision, Clinical Modification codes. We classified the reasons for readmissions as either procedure- or medical-related. Cost-to-charge ratios supplied with the Nationwide Readmissions Database were used to compute the individual per-patient cost of 90-day readmissions as a continuous variable in separate general linear models for THA and TKA. Payer, patient, clinical, and hospital factors were treated as covariates. We estimated the national burden of readmissions by payer and by the reason for readmission.

Results

The national rates of 30- and 90-day readmissions after THA were 4% (95% confidence interval [CI], 4.2%–4.5%) and 8% (95% CI, 7.5%–8.1%), respectively. The national rates of 30- and 90-day readmissions after primary TKA were 4% (95% CI, 3.8%–4.0%) and 7% (95% CI, 6.8%–7.2%), respectively. The five most important variables responsible for the cost of 90-day THA readmissions (in rank order, based on the Type III F-statistic, p < 0.001) were length of stay (LOS), all patient-refined diagnosis-related group (APR DRG) severity, type of readmission (that is, medical- versus procedure-related), hospital ownership, and age. Likewise, the five most important variables responsible for the cost of 90-day TKA readmissions were LOS, APR DRG severity, gender, hospital procedure volume, and hospital ownership. After adjusting for covariates, mean 90-day readmission costs reimbursed by private insurance were, on average, USD 1324 and USD 1372 greater than Medicare (p < 0.001) for THA and TKA, respectively. In the 90 days after TJA, two-thirds of the total annual readmission costs were covered by Medicare. In 90 days after THA, more readmissions were still associated with procedure-related complications, including infections, dislocations, and periprosthetic fractures, which in aggregate account for 59% (95% CI, 59.1%–59.6%) of the total readmission costs to the US healthcare system. For TKA, 49% of the total readmission cost (95% CI, 48.8%–49.6%) in 90 days for the United States was associated with procedure issues, most notably including infections.

Conclusions

Hospital readmissions up to 90 days after TJA represent a massive economic burden on the US healthcare system. Approximately half of the total annual economic burden for readmissions in the United States is medical and unrelated to the joint replacement procedure and half is related to procedural complications.

Clinical Relevance

This national study underscores LOS during readmission as a primary cost driver, suggesting that hospitals and doctors further optimize, to the extent possible, the clinical pathways for the hospitalization of readmitted patients. Because patients readmitted as a result of infection, dislocation, and periprosthetic fractures are the most costly types of readmissions, efforts to reduce the LOS for these types of readmissions will have the greatest impact on their economic burden. Additional clinical research is needed to determine the extent to which, if any, the LOS during readmissions can be reduced without sacrificing quality or access of care.

     

Hormone Receptors,Her-2/Neu and Chromosomal Aberrations in Breast Cancer

Background

There is a great deal of disparity in the incidence of breast cancer in rural and urban India on one hand and between India and Western population on the other.

Methods

We analysed steroid receptor status in cases of breast cancer in a small sample of patients in armed forces. Infiltrating duct carcinomas of breast recorded histologically in mastectomy specimens in last two years were accessioned in the present study with reference to patient and tumour characteristics.

Result

In contrast to the higher rates reported in western literature, only 33 % of the tumours expressed estrogen receptors (ER) and progesterone receptors (PR), of which 24% were ER positive and 30% PR positive. Negative steroid receptor status did not correlate with presence or absence of metastatic nodes, however it was predominant amongst the high grade infiltrating duct carcinomas in this study. Necrosis and lymphovascular invasion demonstrated an inverse relationship with the ER/ PR reactivity. 70% of the node positive cases expressed Her –2/ Neu, reflecting a higher immunoreactivity in this subset of patients. Aneusomy for chromosomes 1, 11 and 17 was common in node positive cases.

Conclusion

Evaluation of chromosomal aberrations by Fluorescent In Situ Hybridization (FISH) technique correlates well with traditional histological parameters.Key Words: Breast carcinoma, Hormone receptors, Her –2/ Neu