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PubMed1976

B cell tolerance induced by polymeric antigens. II. Effects of tolerance on hapten-binding lymphocyte levels in primary and secondary antibody responses.

Tolerogenic doses of hapten [2,4-dinitrophenyl (DNP)]-coupled type 3 pneumococcal polysaccharide (DNP-lys2.5-S3) totally abolished the anti-DNP rosette-forming cell (RFC) response to primary immunization with DNP-hemocyanin in mice, while lightly substituted antigen (DNP-lys0.6-S3) had little effect. Both antigens suppressed secondary anti-DNP RFC responses to DNP-KLH. Limiting doses of DNP-lys-S3 preferentially suppressed antibody-secreting cell levels, and had less effect on RFC. DNP-lys2.5-S3 was 500--1000-fold more potent in "blockading" primary RFC in vitro than DNP-lys0.6-S3, whereas both antigens were equally effective in blocking secondary RFC. These results suggest that the sensitivity of primed B lymphocytes to inactivation by DNP-lys-S3 is related to their high avidity for antigen. Furthermore, this appears to be largely due to a high density of immunoglobulin receptors on primed cells since the affinities of primary and secondary RFC for monovalent hapten were indistinguishable. Treatment of primarily immunized mice with DNP-lys2.5-S3 2 h before assay abolished 90% of RFC. Therefore, the reduction in RFC levels in tolerant mice may be due to cellular blockade by persisting tolerogen. However, it seems unlikely that simple blockade of antigen-reactive cells is the sole mechanism operative in this system.

Animals↗

PubMed2026

Mechanistic diversity of clamp loading at small DNA gaps.

DNA sliding clamps, including PCNA (proliferating cell nuclear antigen) and the 9-1-1 (RAD9-RAD1-HUS1 in humans) complex, are ring-shaped protein complexes that encircle DNA and serve as central interaction platforms in DNA replication, repair, and checkpoint signaling. While clamp loading at canonical primer-template junctions by AAA+ (ATPases associated with diverse cellular activities) clamp loaders is well established, how clamps are loaded onto physiologically relevant but geometrically constrained DNA intermediates, such as nicks and single-stranded gaps, has remained unclear. Recent cryo-electron microscopy studies reveal that clamp loaders have evolved distinct strategies to overcome these constraints and to specialize for different genomic contexts. At gapped DNA, the eukaryotic clamp loader RFC (replication factor C) engages both 3'- and 5'-recessed DNA ends and can locally unwind DNA, enabling PCNA loading across a wide range of gap sizes. In contrast, the bacterial DnaX clamp loader lacks a 5'-DNA-binding site and does not unwind DNA, instead loading the β-clamp at small gaps (<6 nt) by sharply bending DNA. The checkpoint clamp loader Rad24-RFC (RAD17-RFC in humans) similarly lacks DNA unwinding activity, restricting loading of 9-1-1 clamp to larger gaps (≥6 nt). In a distinct specialization, Ctf18-RFC interacts with the leading-strand DNA polymerase ε, positioning it as a dedicated loader for leading-strand synthesis, whereas Elg1-RFC (ATAD5-RFC in humans) excludes DNA from its chamber and functions as a PCNA unloader. Together, these mechanisms illustrate how clamp loaders are diversified to accommodate DNA structure and replisome context, ensuring coordinated control of genome replication and maintenance.

9-1-1 clamp↗

PubMed1976

Assessment of the functional activity of human lymphocytes in malignant disease by the local graft-versus-host reaction in rats and the T rosette-forming cell test.

The local graft-versus-host reaction (GVHR) and the spontaneous rosette-forming cell (RFC) test were used to test the functional activity of lymphocytes in blood obtained from twenty normal donors and thirty patients with malignant tumours. The lymphocytes of all twenty control donors gave a positive GVHR and had normal RFC values. In twenty-two of the thirty patients with malignancies the GVHR was negative; of these, only five had low RFC values. In the remaining eight patients the GVHR was positive and RFC values were normal. It is concluded that the local GVHR constitutes a sensitive test for measurement of the functional activity of lymphocytes, while the RFC test can only be used as a quantitative test.

Animals↗

PubMed2026

Smart crutch tipsTM real-time feedback improves adherence to partial weight-bearing protocols following lower extremity fracture surgery: a pilot study.

PURPOSE: To evaluate the effectiveness of Smart Crutch Tips™ in promoting adherence to prescribed early partial weight-bearing (PWB) protocols and to assess patient experience with real-time weight-bearing feedback during home rehabilitation. METHODS: Twenty patients with lower extremity fractures prescribed partial weight-bearing (PWB) were randomized into two groups utilizing real-time feedback crutch tips (RFC). The Intervention group (n = 10) used Smart Crutch Tips™ (ComeBack Mobility Inc., Kiev, Ukraine), which provided real-time feedback on weight-bearing compliance. The Control group (n = 10) also used crutches equipped with Smart Crutch Tips™, but notifications were disabled, allowing passive data collection without patient feedback. Weight-bearing compliance was defined as the percentage of steps within ± 10% of the prescribed target. Secondary outcomes included patient satisfaction and device usability assessed using the System Usability Scale (SUS) throughout the home rehabilitation period. RESULTS: The Intervention group demonstrated significantly greater PWB compliance compared with the Control group (73.6% vs. 21.1%, p < 0.01). The Intervention group reported a mean SUS score of 84.25, indicating excellent usability. No device-related complications or adverse events were observed. Patient satisfaction, assessed through video interviews, was high, with participants in the Intervention group expressing a strong willingness to recommend the device to others with similar injuries and rating it 10 out of 10. CONCLUSION: Smart Crutch Tips™ significantly improved adherence to prescribed partial weight-bearing protocols following lower extremity fracture surgery. The RFC device demonstrated high usability, excellent patient satisfaction, and no device-related complications during home rehabilitation throughout the study period.

Humans↗

PubMed1977

Demonstration and characterization of a serum factor produced by activated T cells.

Spleen rosette forming cells (RFC) from adult thymectomized mice have a low sensitivity to inhibition by anitheta serum (AOS) and azathioprine (AZ) in comparison with normal spleen or thymus RFC. Thymus extracts and normal mouse serum (but not spleen extracts or thymectomized mouse serum) correct this abnormality after a 30 min in vitro incubation with spleen cells. We report here the existence of a serum factor produced in allogeneic reactions with the same activity on rosettes as thymic factor (TF). This 'allogeneic' factor (AF) is detectable in mice undergoing a graft versus host reaction (GVHR), rejecting skin allografts or allogeneic cells or responding to thymus-dependent antigens such as heterologous red blood cells or BSA. The T-cell origin of AF is indicated by AF presence in nude mice submitted to the same allogeneic stimuli as listed above and in normal mice injected with PVP or LPS. AF is distinct from the thymic factor as shown by differences in electric charge. Moreover, in contrast with TF there is no specific high molecular weight inhibitor of AF. Preliminary biochemical studies indicate that AF is probably a peptide of low molecular weight (greater than 5000 daltons). Its target cell is probably a T-cell precursor.

Animals↗

PubMed2025

The DNA replication checkpoint limits Okazaki fragment accumulation to protect and restart stalled forks.

Understanding how DNA replication forks stall and restart and how the DNA replication checkpoint prevents irreversible fork collapse in molecular detail are crucial for understanding how cells maintain stable genomes and how they prevent the genetic instability that drives cancer. Here, we describe the reconstitution of fork stalling and restart with purified budding yeast proteins. After nucleotide depletion, leading-strand DNA synthesis quickly stops but CMG helicase continues to unwind, and Okazaki fragments continue to initiate on the lagging strand. Incomplete Okazaki fragments sequester PCNA, RFC, and DNA polymerases δ and ε, which prevents normal DNA synthesis restart and exposes nascent DNA to nuclease attack. The DNA replication checkpoint restrains fork progression, which limits this sequestration, protecting stalled forks from collapse and ensuring restart.

DNA Replication↗