Increased Frequency of Mutations in the <i>hprt</i> Gene of T Cells Isolated from Patients with Anti-U1-70kD-Autoantibody-Positive Connective Tissue Disease

Abstract

Mixed connective tissue disease (MCTD) is characterized by the presence of high titers of anti-U1-70kD autoantibodies which are the result of substantial B cell activation. The hprt gene encodes the constitutively expressed enzyme hypoxanthine-guanine phosphoribosyl transferase which is active in the purine salvage pathway. Rapidly dividing cells randomly accumulate gene mutations, including mutations in the hprt gene. These mutations may be used to identify activated cells. If activated T cells play a role in the pathogenesis of MCTD, an increased frequency of mutations in the hprt gene might be expected among T cells isolated from such patients. To examine this hypothesis, we isolated and cloned T cells from 10 anti-U1-70kD-autoantibody-positive MCTD patients and determined the precursor frequencies of cells possessing mutations in hprt by comparing the frequency of cells grown in the presence and absence of the purine analogue 6-thioguanine. We found that the frequency of 6-thioguanine-resistant hprt-negative T cells was significantly increased among MCTD patients (mean 566/10⁶; range 122–2,845/10⁶) versus age- and sex-matched controls (mean 42/106; range 21–78/10⁶; p < 0.003). These results demonstrate that there is an increase in the measured mutant frequency of T cells from MCTD patients. Such T cells may play a role in the pathogenesis of this disease.

Assessment of the Frequency of Mutant (hprt-) T Lymphocytes from Peripheral Blood of Patients with Hashimoto's Thyroiditis

“…The observed variance among individual CE estimates under selection or nonselection conditions is also not unusual (13) as it is mainly influenced by the frequency of T cells in the sample. Our results, however, are in contrast to findings for other T-cell-mediated autoimmune diseases such as multiple sclerosis (15,17), type 1 diabetes mellitus (19), rheumatoid arthritis (20), or systemic autoimmune syndromes (17,21,22,24,25), because they do not support a significant difference between the MF values from healthy controls and the patient groups. The reasons for this discrepancy are unclear but the following interpretations are being proposed to …”

“…Under these premises, the mean frequency of mutant (hprt 2 ) T cells in the HT patient population is expected to be higher than the mean frequency of hprt 2 T cells in healthy subjects, because there should be a stronger proliferative response to thyroid antigens, (i.e., more actively dividing T cells) in the first group. This approach has been analogously used to study the frequency, phenotype, or function of peripheral blood mutant T cells in other autoimmune diseases such as multiple sclerosis (15)(16)(17)(18), type 1 diabetes mellitus (19), rheumatoid arthritis (20), systemic lupus erythematosus (21)(22)(23), mixed connective tissue disease (24), and scleroderma (25).…”

HPRT mutations, TCR gene rearrangements, and HTLV‐1 integration sites define in vivo T‐cell clonal lineages

“…Studies have now been reported for multiple sclerosis [Allegretta et al, 1990], Guillain-Barre syndrome [Van den Berg et al, 1995], systemic lupus erythematosus Wood et al, 1994;Theocharis et al, 1995], systemic sclerosis [Sfikakis et al, 1994], mixed connective tissue disease [Holyst et al, 1994], rheumatoid arthritis [Cannons et al, 1998], type 1 diabetes mellitus [Falta et al, 1999[Falta et al, , 2000, paroxysmal nocturnal hemoglobinuria [Chen et al, 2004], and malignant melanoma , with results consistent with the basic hypothesis [Albertini, 2001]. On a practical level, it has been found that sudden and dramatic elevations of HPRT MFs in cardiac transplant recipients have been reliable noninvasive indicators of rejection episodes [Ansari et al, 1995].…”

Selection ofhprtMutant T Cells as Surrogates for Dividing Cells Reveals a Restricted T Cell Receptor BV Repertoire in Insulin-Dependent Diabetes Mellitus

“…Hprt mutations have also been used to estimate the recycling rate of T cells in CD4 ϩ depleted, HIV-infected patients (56). In autoimmune diseases, elevated hprt MFs have been found in the T cells of patients with systemic lupus erythematosus (45)(46)(47), systemic sclerosis (48), mixed connective tissue disease (49), multiple sclerosis (42,50) and other demyelinating disorders (51). Several reports have demonstrated a correlation between MF and parameters of disease status, duration, and severity (46,50,51).…”