In recent years, overlap in biologic and morphologic features has been identified between classic Hodgkin lymphoma (cHL) and B-cell non-Hodgkin lymphoma. Nevertheless, the therapeutic approaches for these diseases remain different. We undertook a study of "mediastinal gray zone lymphomas" (MGZL), with features transitional between cHL nodular sclerosis (NS) and primary mediastinal large B-cell lymphoma (MLBCL) to better understand the morphologic and immunophenotypic spectrum of such cases. Twenty-one MGZL cases were identified over a 20-year period. We also studied 6 cases of composite or synchronous lymphoma with two distinct components at the same time (cHL-NS and MLBCL) and 9 sequential cases with MLBCL and cHL-NS at different times. All patients had a large mediastinal mass. Immunohistochemical studies focused on markers known to discriminate between cHL and MLBCL, including B-cell transcription factors. VJ-PCR was performed in 8 cases to look at clonality of the immunoglobulin heavy chain gene (IgH). Of the gray zone cases, 11 had morphology reminiscent of cHL-NS, but with unusual features, including a large number of mononuclear variants, diminished inflammatory background, absence of classic Hodgkin phenotype, and strong CD20 expression (11 of 11). Ten cases had morphology of MLBCL, but with admixed Hodgkin/Reed-Sternberg and lacunar cells, absent (3 of 10) or weak (7 of 10) CD20 expression, and positivity for CD15 in 7 cases. B-cell transcription factor expression in the gray zone cases more closely resembled MLBCL than cHL with expression of Pax5, Oct2, and BOB.1 in all but 1 case studied (14 of 15). MAL staining was found in 7 of 10 MGZL, and in at least one component of 6 of 7 evaluable composite or sequential MLBCL/cHL cases. Two cases of sequential lymphoma showed rearrangements of the IgH gene of identical size: one in which MLBCL was the first diagnosis and one in which MLBCL was diagnosed at relapse, indicating clonal identity for the two components of cHL and MLBCL. There is accumulating evidence that MLBCL and cHL are related entities. Further support for a relationship between MLBCL and cHL-NS is provided by composite and metachronous lymphomas in the same patient, as well as the existence of MGZL with transitional morphology and phenotype.

Combinatorial association of different lipid species generates microheterogeneity in biological membranes. The association of glycosphingolipids with cholesterol forms membrane microdomains – lipid rafts – that are involved in specialised pathways of protein/lipid transport and signalling. Lipid rafts are normally dispersed in cellular membranes and appear to require specialised machinery to reorganise them to operate. Caveolin-1 and MAL are members of two different protein families involved in reorganisation of lipid rafts for signalling and/or intracellular transport in epithelial cells. T cell activation induces a rapid compartmentalisation of signalling machinery into reorganised rafts that are used as platforms for the assembly of the signalling complex. Costimulatory molecules participate in this process by providing signals that mobilise raft lipids and proteins, and remodel the cytoskeleton to the contact site. As in epithelial cells, rafts are used also as vesicular carriers for membrane trafficking in T lymphocytes. Furthermore, there are potential similarities between the specialised protein machinery underlying raft-mediated processes in T lymphocytes and polarised epithelial cells.

The MAL (MAL/VIP17) proteolipid is a nonglycosylated integral membrane protein expressed in a restricted pattern of cell types, including T lymphocytes, myelin-forming cells, and polarized epithelial cells. Transport of the influenza virus hemagglutinin (HA) to the apical surface of epithelial Madin-Darby canine kidney (MDCK) cells appears to be mediated by a pathway involving glycolipid- and cholesterol- enriched membranes (GEMs). In MDCK cells, MAL has been proposed previously as being an element of the protein machinery for the GEM-dependent apical transport pathway. Using an antisense oligonucleotide-based strategy and a newly generated monoclonal antibody to canine MAL, herein we have approached the effect of MAL depletion on HA transport in MDCK cells. We have found that MAL depletion diminishes the presence of HA in GEMs, reduces the rate of HA transport to the cell surface, inhibits the delivery of HA to the apical surface, and produces partial missorting of HA to the basolateral membrane. These effects were corrected by ectopic expression of MAL in MDCK cells whose endogenous MAL protein was depleted. Our results indicate that MAL is necessary for both normal apical transport and accurate sorting of HA.

How the vesicular traffic of signaling molecules contributes to T cell receptor (TCR) signal transduction at the immunological synapse remains poorly understood. In this study, we show that the protein tyrosine kinase Lck, the TCRζ subunit, and the adapter LAT traffic through distinct exocytic compartments, which are released at the immunological synapse in a differentially regulated manner. Lck vesicular release depends on MAL protein. Synaptic Lck, in turn, conditions the calcium- and synaptotagmin-7–dependent fusion of LAT and TCRζ containing vesicles. Fusion of vesicles containing TCRζ and LAT at the synaptic membrane determines not only the nanoscale organization of phosphorylated TCRζ, ZAP70, LAT, and SLP76 clusters but also the presence of phosphorylated LAT and SLP76 in interacting signaling nanoterritories. This mechanism is required for priming IL-2 and IFN-γ production and may contribute to fine-tuning T cell activation breadth in response to different stimulatory conditions.