Carbonic anhydrase (CA) catalyzes the reversible hydration of CO(2). CA is expressed in most segments of the kidney. CAII and CAIV predominate in human and rabbit kidneys; in rodent kidneys, CAXII, and CAXIV are also present. CAIX is expressed by renal cell carcinoma (RCC). Most of these isoforms, except for rodent CAIV, have high turnover rates. CAII is a cytoplasmic enzyme, whereas the others are membrane-associated; CAIV is anchored by glycosylphosphatidylinositol linkage. Membrane polarity is apical for CAXIV, basolateral for CAXII, and apical and basolateral for CAIV. Luminal membrane CAs facilitate the dehydration of carbonic acid (H(2)CO(3)) that is formed when secreted protons combine with filtered bicarbonate. Basolateral CA enhances the efflux of bicarbonate via dehydration of H(2)CO(3). CAII and CAIV can associate with bicarbonate transporters (e.g., AE1, kNBC1, NBC3, and SCL26A6), and proton antiporter, NHE1 in a membrane protein complex called a transport metabolon. CAXII and CAXIV may also be associated with transporters in normal kidney and CAIX in RCCs. The multiplicity of CAs implicates their importance in acid-base and other solute transport along the nephron. For example, CAII on the cytoplasmic face and CAIV on the extracellular surface provide the 'push' and 'pull' for bicarbonate transport by supplying and dissipating substrate respectively.

The Na + concentration of the intracellular milieu is very low compared with the extracellular medium. Transport of Na + along this gradient is used to fuel secondary transport of many solutes, and thus plays a major role for most cell functions including the control of cell volume and resting membrane potential. Because of a continuous leak, Na + has to be permanently removed from the intracellular milieu, a process that is thought to be exclusively mediated by the Na + /K + -ATPase in animal cells. Here, we show that intercalated cells of the mouse kidney are an exception to this general rule. By an approach combining two-photon imaging of isolated renal tubules, physiological studies, and genetically engineered animals, we demonstrate that inhibition of the H + vacuolar-type ATPase (V-ATPase) caused drastic cell swelling and depolarization, and also inhibited the NaCl absorption pathway that we recently discovered in intercalated cells. In contrast, pharmacological blockade of the Na + /K + -ATPase had no effects. Basolateral NaCl exit from β-intercalated cells was independent of the Na + /K + -ATPase but critically relied on the presence of the basolateral ion transporter anion exchanger 4. We conclude that not all animal cells critically rely on the sodium pump as the unique bioenergizer, but can be replaced by the H + V-ATPase in renal intercalated cells. This concept is likely to apply to other animal cell types characterized by plasma membrane expression of the H + V-ATPase.

Although resting B cells as APC are tolerogenic for naive T cells in vivo, we show here that they can provide all the costimulatory signals necessary for naive T cell proliferation in vivo and in vitro. In the absence of an activating signal through the B cell Ag receptor, T cell proliferation after Ag recognition on resting B cells depends on CD40 expression on the B cells, implying that naive T cells use the membrane-bound cytokine, CD40 ligand (CD154), to induce the costimulatory signals that they need. Induction of B7-1 (CD80) and increased or sustained expression of CD44H, ICAM-1 (CD54), and B7-2 (CD86) are dependent on the interaction of CD40 ligand with CD40. Transient expression (12 h) of B7-2 is T cell- and peptide Ag-dependent, but CD40-independent. Only sustained (≥24 h) expression of B7-2 and perhaps increased expression of ICAM-1 could be shown to be functionally important in this system. T cells cultured with CD40-deficient B cells and peptide remain about as responsive as fresh naive cells upon secondary culture with whole splenic APC. Therefore, B cells, and perhaps other APC, may be tolerogenic not because they fail to provide sufficient costimulation for T cell proliferation, but because they are deficient in some later functions necessary for a productive T cell response.

It is well known that acid/base disturbances modulate proton/bicarbonate transport in the cortical collecting duct. To study the adaptation further we measured the effect of three days of acidosis followed by the rapid recovery from this acidosis on the number and type of intercalated cells in the rabbit cortical collecting duct. Immunofluorescence was used to determine the expression of apical pendrin in β-intercalated cells and the basolateral anion exchanger (AE1) in α-intercalated cells. Acidosis resulted in decreased bicarbonate and increased proton secretion, which correlated with reduced pendrin expression and the number of pendrin-positive cells, as well as decreased pendrin mRNA and protein abundance in this nephron segment. There was a concomitant increase of basolateral AE1 and α-cell number. Intercalated cell proliferation did not seem to play a role in the adaptation to acidosis. Alkali loading for 6–20 h after acidosis doubled the bicarbonate secretory flux and reduced proton secretion. Pendrin and AE1 expression patterns returned to control levels, demonstrating that adaptive changes by intercalated cells are rapidly reversible. Thus, regulation of intercalated cell anion exchanger expression and distribution plays a key role in adaptation of the cortical collecting duct to perturbations of acid/base.

SummlryWe have examined the contributions of Interleukin 4 (Ib4), IL-5, and other stimuli to the expression of Immunoglobulin G1 (IgG1) and IgE in murine B lymphoblasts activated with anti-Ig. The combination of IL-4 and -5 induced B lymphobhsts to proliferate and to secrete IgM and IgG1. However, an additional stimulus was required along with II:4 and -5 for induction of IgE secretion. This stimulus was provided by lipopolysaccharides (LPS) or cytokines produced by TC-1 or EL4 cells. In the absence of II.-5, exceptionally high concentrations of II.-4 (>1,000 U/ml) were required to elicit IgG1 and IgE secretion from B lymphoblasts cultured with either LPS or TC-l-conditioned media (CM). To investigate regulation of expression of 3'1 and ~ genes by I1,4, -5, and LPS, the requirements for induction of 3'1 and e germline and productive transcripts were examined. Germline 3'1, but not e, transcripts were detected in RNA from B lymphoblasts treated with I1,4 and -5 for 48 h. In contrast, both germline 3'1 and e transcripts could be detected in B lymphoblasts cultured with I1,4 and LPS, and steady state levels of germline 3'1 transcripts were four-to sevenfold higher in blasts cultured with LPS and Ib4, compared with blasts cultured with I1,4 and -5. LPS enhanced steady state levels of germline transcripts induced by I1,4, but LPS did not promote substantial accumulation of productive 3'1 and e transcripts. In contrast, I1,5 did not affect steady state levels of germline transcripts stimulated by I1,4, but did markedly increase levels of productive 3'1 and e transcripts. Thus, lymphokines regulate two distinct events in isotype switching: induction of germline transcripts (I1,4), and production of VDJ-C3'I and VDJ-Ce mRNA (I1,5), which leads to secretion of IgG1 and IgE.