Enhancing effect of murine anti-idiotypic serum on the proliferative response specific for poly(LTyr,LGlu)-poly(DLAla)–poly (LLys) [(T,G)-A–L]

Murine anti-idiotypic serum against C3H.SW anti-poly(LTyr, LGlu)-poly(DLAla)–poly(LLys) [(T, G)-A–L] antibodies was elicited in C57BL/6 mice. The effect of the anti-idiotypes on the proliferation of primed lymph node cells was investigated. The anti-idiotypic serum stimulated the proliferative response of the (T, G)-A–L-specific lymph node cells as well as of nylon wool-enriched T cells. In the presence of suboptimal doses of (T, G)-A–L, the addition of the anti-idiotypes enhanced the proliferation to the levels obtained with the optimal dose of (T, G)-A–L itself. These results suggest the existence of shared idiotypic determinants between antibodies and the (T, G)-A–L-specific proliferative T cells.     

Characteristics of a poly(LTyr,LGlu)-poly(DLAla)--poly(LLys)-specific helper factor derived from a T cell hybridoma

A poly(LTyr,LGlu)-poly(DLAla)-poly(LLys) [(T,G)-A-L]-specific helper T cell hybridoma, R-9, was established by fusing activated T cells, enriched for specific helper activity by incubation on antigen-pulsed splenic adherent cells, with the BW5147 thymoma line. Supernatants of this hybrid line or its active clones as well as ascitic fluids, aspirated from the peritoneum of mice injected with the hybridoma cells, manifest a(T,G)-A-L-specific helper T cell-replacing activity as assayed in an in vitro hapten-carrier antibody production system. The factor activity can be specifically absorbed on and eluted from columns of immobilized (T,G)-A–L, antibodies directed against the variable region of the immunoglobulin heavy chain and an antiserum directed against (T,G)-A-L-specific idiotypes. Although the R-9 factor and anti-(T,G)-A-L antibodies share V region gene-encoded determinants, they differ in their fine antigenic specificity, as determined by cross-reactivity with closely related synthetic polypeptide immunogens. Whereas anti-(T,G)-A–L antibodies bind equally well to the L-phenylalanine- or L-histidine-containing polypeptides (Phe,G)-A-L and (H,G)-A-L, to poly(LGlu)-poly(DLAla)-poly(LLys) (G-A–L) and poly(Tyr,Glu)-poly(LPro)-poly(LLys) [(T,G)-Pro–L], the R-9 factor was completely absorbed on columns of (Phe,G)-A-L and G-A-L and only partially absorbed on immobilized (T,G)-Pro–L and (H,G)-A-L. In addition to V region gene products, the R-9 factor also bears major histocompatibility complex (MHC) determinants, which were found to map to the left end of the I^b region. Combination of inactive effluents obtained after absorption of the R-9 factor on columns of anti-H-2^b and anti-Id antibodies resulted in active factor preparations, suggesting that although V region gene and MHC products are linked together in the active part of the factor, these moieties are located on different polypeptide chains which may associate in solution.     

Elicitation and detection of in vitro H-2-linked Ir gene regulated antibody responses to poly(LTyr, Glu)-poly(DLAla)--poly(LLys)

A microculture system is described in which secreted antibody responses to the synthetic polypeptide (T,G)-A--L were obtained in vitro. Responses were highly reproducible, antigen-dependent, antigen-specific, and under H-2-linked Ir gene control. Critical elements in the system include the schedule of in vivo antigen-priming, removal of the stimulating antigen after 3 days of culture, and a sensitive detection system (double-antibody ELISA). This system should be useful in the analysis of the mechanism of action of Ir genes as well as the mechanisms by which anti-idiotype antibodies modulate immune responses.     

Fine specificity and idiotypic expression of monoclonal antibodies directed against poly(Tyr,Glu)-poly(DLAla)--poly(Lys) and its ordered analogue (Tyr-Tyr-Glu-Glu)-poly(DLAla)--poly(Lys).

In order to study the repertoire of poly(Tyr,Glu)-poly(DLAla)--poly(Lys) [(T,G)-A--L] specific antibodies, monoclonal antibodies were prepared by fusing myeloma cells with spleen cells from C3H.SW mice immunized with (T,G)-A--L and boosted with (Tyr-Tyr-Glu-Glu)-poly(DLAla)--poly(Lys)](T-T-G-G)-A--L]. Eleven clones which secreted homogeneous antibodies were obtained. In general, two families of monoclonal antibodies were detected: those which bind exclusively (T-T-G-G)-A--L and those which bind both (T-T-G-G)-A--L and (T,G)-A--L. Analysis for idiotypic expression revealed that only two antibodies (clones no. 103 and 160), which were found to be similar in their fine specificity, cross-reacted with antibodies against the major idiotypes of (T,G)A--L specific antibodies. Guinea-pig antibodies against clone no. 160 reacted with the polyclonal (T,G)-A--L specific antibodies, whereas antibodies against 103 monoclonal antibodies did not react with C3H.SW anti-(T,G)-A--L antibodies, but did cross-react with four other monoclonal antibodies. It appears that the idiotypic determinants expressed on polyclonal (T,G)-A--L specific antibodies are heterogeneous, and consist of at least two serologically different idiotypes detected by clones no. 103 and 160.     

SLC26A2 disease spectrum in Sweden – high frequency of recessive multiple epiphyseal dysplasia (rMED)

Diastrophic dysplasia (DTD) is an autosomal recessive skeletal dysplasia caused by SLC26A2 mutations. Clinical features include short stature, joint contractures, spinal deformities, and cleft palate. SLC26A2 mutations also result in other skeletal dysplasias, including the milder recessive multiple epiphyseal dysplasia (rMED). DTD is overrepresented in Finland and we speculated that this may have influenced the prevalence and spectrum of SLC26A2‐related skeletal conditions also in Sweden. We reviewed the patient registry at Department of Clinical Genetics, Karolinska University Hospital, Stockholm to identify subjects with SLC26A2 mutations. Seven patients from six families were identified; clinical data were available for six patients. All but one patient had one or two copies of the Finnish SLC26A2 founder mutation IVS1+2T>C. Arg279Trp mutation was present in compound heterozygous form in five patients with phenotypes consistent with rMED. Their heights ranged from ?2.6 to ?1.4 standard deviation units below normal mean and radiographic features included generalised epiphyseal dysplasia and double‐layered patellae. Two rMED patients had hypoplastic C2 and cervical kyphosis, a severe manifestation previously described only in DTD. Our study confirms a high prevalence of rMED in Sweden and expands the phenotypic manifestations of rMED.