Abnormal development of secondary lymphoid tissues in lymphotoxin β-deficient mice

The tumor necrosis factor (TNF) family cytokines lymphotoxin (LT) α and LTβ form heterotrimers that are expressed on the surface of activated lymphocytes and natural killer cells; LTα homotrimers can be secreted as well. Mice with a disrupted LTα gene lack lymph nodes (LN), Peyer’s patches (PP), and follicular dendritic cell (FDC) networks and reveal profound defects of the splenic architecture. However, it is unclear which of these abnormalities is the result of the absence in LTα homotrimers or LTαβ heterotrimers. To distinguish between these two possibilities, a mouse strain deficient in LTβ was created employing Cre/loxP-mediated gene targeting. Mice deficient in LTβ reveal severe defects in organogenesis of the lymphoid system similar to those of LTα−/− mice, except that mesenteric and cervical LN are present in most LTβ-deficient mice. Both LTβ- and LTα-deficient mice show significant lymphocytosis in the circulation and peritoneal cavity and lymphocytic infiltrations in lungs and liver. After immunization, PNA-positive B cell clusters were detected in the splenic white pulp of LTβ-deficient mice, but FDC networks were severely underdeveloped. Collectively, these results indicate that LTα can signal independently from LTβ in the formation of PNA-positive foci in the spleen, and especially in the development of mesenteric and cervical LN.

Reduction of HIV-1 in blood and lymph nodes following potent antiretroviral therapy and the virologic correlates of treatment failure

Potent antiretroviral therapy can reduce plasma HIV RNA levels below the threshold of detection for periods of a year or more. The magnitude of HIV RNA reduction in the lymphoid tissue in patients with suppression of HIV RNA levels in plasma beyond 6 months has not been determined. We evaluated levels of HIV RNA and DNA and characterized resistance mutations in blood and inguinal lymph node biopsies obtained from 10 HIV-infected subjects who received 36–52 weeks of indinavir (IDV)/zidovudine (ZDV)/lamivudine (3TC), IDV, or ZDV/3TC. After 1 year of therapy, viral RNA levels in LN of individuals remained detectable but were log10 = 4 lower than in subjects on the triple drug regimen with interruption of therapy or in those treated with ZDV/3TC alone, who had viral loads in their lymph nodes indistinguishable from those expected for untreated patients. In all cases viral DNA remained detectable in lymph nodes and peripheral blood mononuclear cells (PBMC). When plasma virus suppression was incomplete, lymph node and PBMC cultures were positive and drug resistance developed. These studies indicate that pronounced and sustained suppression of plasma viremia by a potent antiretroviral combination is associated with low HIV RNA levels in the lymph nodes 1 year after treatment. Conversely, the persistence of even modest levels of plasma virus after 1 year of treatment reflects ongoing viral replication, the emergence of drug resistance, and the maintenance of high burdens of virus in the lymph nodes.

Integrin α6Aβ1 Induces CD81-dependent Cell Motility without Engaging the Extracellular Matrix Migration Substrate

It is well established that integrins and extracellular matrix (ECM) play key roles in cell migration, but the underlying mechanisms are poorly defined. We describe a novel mechanism whereby the integrin α6β1, a laminin receptor, can affect cell motility and induce migration onto ECM substrates with which it is not engaged. By using DNA-mediated gene transfer, we expressed the human integrin subunit α6A in murine embryonic stem (ES) cells. ES cells expressing α6A (ES6A) at the surface dimerized with endogenous β1, extended numerous filopodia and lamellipodia, and were intensely migratory in haptotactic assays on laminin (LN)-1. Transfected α6A was responsible for these effects, because cells transfected with control vector or α6B, a cytoplasmic domain α6 isoform, displayed compact morphology and no migration, like wild-type ES cells. The ES6A migratory phenotype persisted on fibronectin (Fn) and Ln-5. Adhesion inhibition assays indicated that α6β1 did not contribute detectably to adhesion to these substrates in ES cells. However, anti-α6 antibodies completely blocked migration of ES6A cells on Fn or Ln-5. Control experiments with monensin and anti-ECM antibodies indicated that this inhibition could not be explained by deposition of an α6β1 ligand (e.g., Ln-1) by ES cells. Cross-linking with secondary antibody overcame the inhibitory effect of anti-α6 antibodies, restoring migration or filopodia extension on Fn and Ln-5. Thus, to induce migration in ES cells, α6Aβ1 did not have to engage with an ECM ligand but likely participated in molecular interactions sensitive to anti-α6β1 antibody and mimicked by cross-linking. Antibodies to the tetraspanin CD81 inhibited α6Aβ1-induced migration but had no effect on ES cell adhesion. It is known that CD81 is physically associated with α6β1, therefore our results suggest a mechanism by which interactions between α6Aβ1 and CD81 may up-regulate cell motility, affecting migration mediated by other integrins.

L-selectin can facilitate metastasis to lymph nodes in a transgenic mouse model of carcinogenesis

L-selectin mediates homing of lymphocytes to lymph nodes (LN). Transgenic mice that express rat insulin promoter regulated simian virus 40 Tag (RIP-Tag) develop large, local cancers that metastasize to liver but not LN. To test whether this lack of LN metastases reflects their absence from the circulation, transgenic mice were produced that express Tag (T), L-selectin (L), and Escherichia coli LacZ (Z), in pancreatic β cells. LTZ mice developed insulinomas that specifically had LN metastases; metastasis was blocked by an anti L-selectin mAb. LacZ+ tumor cells from these LN homed to secondary LN upon transfer. These results suggest that the highly vascularized islet carcinomas are shedding tumor cells into the bloodstream, which is a necessary but insufficient condition for metastasis to occur; L-selectin can facilitate homing of such tumor cells to LN, resulting in metastasis.

Significant discrepancies between van't Hoff and calorimetric enthalpies.

In this paper we show that the usual assumption in studies of the temperature variation of equilibrium constants for equilibria of the form A+B <-->AB that a plot of ln K vs. 1/T (K = equilibrium constant, T = temperature in degrees kelvin) is a straight line with slope equal to -delta HvH/R (delta HvH = van't Hoff or apparent enthalpy, R = gas constant) is not valid in many cases. In all the cases considered here, delta HvH is temperature dependent and is significantly different from the true or calorimetrically measured enthalpy, and the respective values for delta Cp are also significantly different.