T-cell hybridomas are powerful tools in studying the fine specificities of antigen recognition by the T-cell receptor (TCR), the structure and genetic basis of the CD3—TCR complex, and the size of the TCR α/β repertoire used in response to various antigens. A technical challenge in establishing T-cell hybridomas is the early identification of antigen-specific ones. We have established a rapid and efficient ELISA method for detecting antigen-specific T-cell hybridomas. Our ELISA technique significantly reduces the time and resources required for the primary screening of antigen-specific T-cell hybrids, eliminates the need of maintaining hundreds of rapidly growing nonspecific clones, and does not require the maintenance of IL-2/IL-4 dependent cell lines such as CTLL-2 or HT-2. In addition, the ELISA technique is designed to detect both types of CD4 T-cells: Th1 and Th2, by using a mixture of anti-IL-2 and anti-IL-4 monoclonal antibodies. Therefore, we believe that our ELISA technique provides a faster, less expensive, and higher throughput screening method for the early identification of antigen-specific T-cell hybridomas than the current bioassays.
Physical resilience, the capacity to respond to and recover from a stressful event, declines with advancing age. Individuals respond differently to physical stressors across their lifespans. While the biological underpinnings of resilience remain unclear, a plausible determinant is the capacity of an individual’s cellular and molecular levels to return to homeostasis after a physical challenge. Impaired resilience may not only be a consequence of aging but could also be a contributing factor to the aging process. Therefore, resilience at relatively younger ages could be predictive of future health and lifespan. By utilizing standardized physical challenges and measuring stress response patterns, the relative resilience of individuals can be quantified and classified. Current preclinical research suggests that several physical stressors could be used to measure resilience in clinical aging studies. A mechanistic understanding of why some individuals are more resilient to physical stressors than others could help identify protective factors and therapeutic ways to promote healthy aging. Keywords: Physical resilience to aging, physical stressors, heterogeneity, stress response patterns, healthy aging, therapeutic resilience
Twenty-three of 37 anti-la McAb reactive with human B cells, as determined by indirect immunocytofluorescence, were shown to be reactive with canine peripheral blood mononuclear cells (PBMC). Using a panel of human B cell lines that differ in their expression of HLA-DR,-DP, and-DQ molecules, it was shown that 15 of these antibodies identify HLA-DR and DP molecules (i.e., broadly reactive), while 22 identify only HLA-DR molecules. Fourteen of the 15 broadly reactive McAb were reactive with canine PBMC while only 9 of the 22 HLA-DR-specific McAb reacted with canine PBMC, suggesting that broadly reactive anti-Ia McAb are much more likely to react with canine cells than narrowly reactive McAb. Ten of the canine reactive McAb that were shown to identify typical Ia bimolecular structures on canine cells using immune precipitation analysis were tested for blocking activity in the canine mixed lymphocyte culture (MLC). All four of the broadly reactive McAb (B1F6, J-70, 9–49, and HB10a) plus two of the six narrowly reactive McAbs (H81.98.21 and H40.164.3) blocked the canine MLC when added to culture wells on day 0, suggesting that inhibition may be related to the specificity of the anti-Ia McAb employed. Since the MLC may reflect cellular interactions occurring during graft-versus-host disease, this assay may be useful for screening functionally relevant broadly reactive McAb in experimental canine bone marrow transplantation studies. These data suggest that the dog may be a useful model to study anti-la immuno-therapy.
Anxiety is a pervasive emotional response that can profoundly impact well-being and cognitive function in both humans and animals. The relationship between anxiety and aging remains complex and multifaceted. To explore this relationship in more detail, an open-field photobeam system was used to quantify anxiety-related behaviors in aging CB6F1 and C57BL/6 male mice and to determine associations with aging phenotypes, including short- and long-term memory, grip strength, rotarod performance, and self-motivated wheel running. Results indicated a heightened anxiety in novel environments with increasing age as evidenced by a preference for peripheral areas during the open-field test. Elevated anxiety levels were not associated with decreased cognitive performance, suggesting that anxiety and cognition operate somewhat independently of each other. A negative correlation was observed between anxiety levels and distance run in the voluntary wheel running assessment, while no associations were seen with grip strength or rotarod performance. These observations contribute to a better understanding of anxiety and its consequences in aging mice, providing insights into potential therapeutic interventions aimed at delaying aging through anxiety management. Keywords : Anxiety, aging, behavioral assessment, cognition, CB6F1 mice, C57BL/6 mice
Tumor progression depends on the support of cells in the microenvironment, and is driven in part by the generation of reactive oxygen species (ROS). ROS can damage DNA, and the repair of damaged DNA is a well-known process involved in tumor initiation and promotion, but the role of DNA repair in tumor progression is not fully understood. In this regard the X-ray cross complementing 1 (XRCC1) protein is known to orchestrate the assembly of repair complexes at sites of DNA single strand breaks either directly or indirectly through repair of damaged bases, largely as the result of ROS-induced damage. XRCC1 polymorphisms have been shown to be associated with increased cancer. It was therefore of interest to investigate the effect of XRCC1 gene mutations on cancer progression. In an attempt to make XRCC1 point mutant mice, we generated a truncated protein (XRCC1tp) by the insertion of a neomycin cassette in intron12 of the XRCC1 gene. This unique finding allowed us to investigate cellular and tumor progression phenotypes in mice associated with expression and function of an altered XRCC1 protein on one allele. XRCC1tp cells showed increased toxicity to MMS, enhanced MMS-induced depletion of NADH suggesting increased PARP activity, and normal functional repair of MMS-induced DNA damage. Six months following treatment with the alkylating carcinogen azoxymethane (AOM) at 10 mg/kg once a week for 6 weeks, XRCC1tp mice had a decrease in average colon tumor volume of 14±3 mm(3) compared to 34±4 mm(3) in WT littermates (p ≤ 0.03, N= 20/genotype). XRCC1tp mice had a 72 per cent decrease in B16 melanoma tumor burden compared to wt littermates. Average tumor volume in transgenic PyMT metastatic breast cancer mice expressing XRCC1tp was 359 cubic mm in PyMT mice expressing XRCC1tp compared to 730 cubic mm in PyMT mice expressing XRCC1wt (p ≤ 0.001, N= 20/genotype). These data suggest that the presence of an XRCC1 truncated protein alters XRCC1 function independent of DNA repair, and is associated with anti-tumor activity.
A recent article by Pyrkov et al [1] suggests that progressive loss of physical resilience to aging coincides with an absolute limit of human lifespan in the range of 120 to 150 years. Furthermore, the authors conclude that end of life is an intrinsic biological property independent of internal or external stress factors
