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We have studied serum cytokine profiles in BALB/c mice after immunization with influenza vaccine alone or combined with the following adjuvants: alum; MF59 emulsion; MF59 containing the muramyl peptide N-acetyl-muramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2- dipalmitoyl-sn-glycero-3-(hydroxyphosphoryloxy)) ethylamide (MTP-PE); MF59 plus the lipid A analogue monophosphoryl lipid A; MF59 plus the Quil A saponin fraction LTC; or LTC alone. Pooled mouse sera were analyzed by ELISA at various times after immunization for IL-1 alpha, IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, IFN-gamma, and TNF-alpha. In naive mice, vaccine alone induced low levels of IL-3 and IL-5 only; vaccine plus alum induced a low IL-6 response as well. The MF59-based adjuvants significantly increased the IL-5 and IL-6 levels, whereas Quil A LTC induced strong IFN-gamma and measurable IL-2 responses, in addition to moderate IL-5 and IL-6. In previously infected mice, MF59 and MF59/MTP-PE were capable of generating IFN-gamma responses, as well as IL-5 and IL-6. All of the cytokine responses were rapid (peaking 3 to 12 h postimmunization) and short lived. In naive mice, the MF59 adjuvants induced serum cytokine profiles that are consistent with a primarily Th2-type response, whereas the Quil A LTC induced cytokines associated with both Th1 and Th2 responses. Ab analyses indicated that, although the adjuvants strongly affected the magnitude of the humoral response, there was no obvious correlation between the cytokine profile observed and the subclasses of Ab induced.

We have studied serum cytokine profiles in BALB/c mice after immunization with influenza vaccine alone or combined with the following adjuvants: alum; MF59 emulsion; MF59 containing the muramyl peptide N-acetyl-muramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2- dipalmitoyl-sn-glycero-3-(hydroxyphosphoryloxy)) ethylamide (MTP-PE); MF59 plus the lipid A analogue monophosphoryl lipid A; MF59 plus the Quil A saponin fraction LTC; or LTC alone. Pooled mouse sera were analyzed by ELISA at various times after immunization for IL-1 alpha, IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, IFN-gamma, and TNF-alpha. In naive mice, vaccine alone induced low levels of IL-3 and IL-5 only; vaccine plus alum induced a low IL-6 response as well. The MF59-based adjuvants significantly increased the IL-5 and IL-6 levels, whereas Quil A LTC induced strong IFN-gamma and measurable IL-2 responses, in addition to moderate IL-5 and IL-6. In previously infected mice, MF59 and MF59/MTP-PE were capable of generating IFN-gamma responses, as well as IL-5 and IL-6. All of the cytokine responses were rapid (peaking 3 to 12 h postimmunization) and short lived. In naive mice, the MF59 adjuvants induced serum cytokine profiles that are consistent with a primarily Th2-type response, whereas the Quil A LTC induced cytokines associated with both Th1 and Th2 responses. Ab analyses indicated that, although the adjuvants strongly affected the magnitude of the humoral response, there was no obvious correlation between the cytokine profile observed and the subclasses of Ab induced.

The synthesis and secretion of proteins during development of the pancreas was analyzed using two-dimensional gel electrophoresis. The pattern of synthesis of the total proteins of the pancreas was found to change very little from 14 to 18 d gestation. In addition, the protein synthetic pattern of the embryonic pancreas was very similar to the protein patterns of several other embryonic tissues (gut, lung, and mesenchyme). Between 18 d gestation and the adult stage, the synthesis of the majority of protein species fades as the synthesis of the secretory (pro)enzymes becomes dominant. Thus, the terminal differentiation of the pancreas appears to involve the dominant expression of a limited set of genes (coding, in part, for the digestive [pro]enzymes) while the pattern of expression of the remaining domain remains relatively unchanged. Many of the secretory (pro)enzymes were identified and their synthesis during development was monitored. The synthesis of several secretory proteins was detected between 15 and 18 d gestation (e.g., amylase and chymotrypsinogen), whereas the synthesis of others was not detected until after 18 d gestation (i.e., trypsinogen, ribonuclease, proelastase, and lipase). Between 18 d gestation and the adult stage, the synthesis of the digestive (pro)enzymes increases to > 90% of pancreatic protein synthesis. The secretion of digestive (pro)enzymes was detected as early as 15 d gestation. The selective release of a second set of proteins was detected in the early embryo. These proteins are not detected in the adult pancreas or in zymogen granules but are also released by several other embryonic tissues. The function of this set of proteins is unknown.

The ability of suspensions of BALB/c cells to catalyze the incorporation of nucleotide sugars into complex polysaccharides has been compared. These cells have previously been characterized for concanavalin A-induced agglutinability, tumorigenicity, and malignancy. All of the cell lines tested catalyze transfer of the sugar moieties of cytosine 5-monophosphate N-acetylneuraminic acid, uridine 5-diphosphate galactose, uridine 5-diphosphate N-acetylgalactosamine, uridine 5-diphosphate N-acetylglucosamine, uridine 5-diphosphate glucose, and guanidine 5-diphosphate monnose into glycoproteins and glycolipids. While some transformed lines exhibit alterations in transferase levels, others cannot be distinguished from normal cells. Normal cells, transformed cells that cause tumors that regress, and transformed cells that cause tumors that kill an immunologically competent host show growth-dependent changes in transferase activities. Determining the ability to catalyze carbohydrate transfer is insufficient for predicting the tumorigenic and malignant properties of a cell line.

Membrane glycolipids, glycoproteins, and surface proteins of normal and transformed BALB/c cell lines have been compared. Several virally and spontaneously transformed cell lines showed differences in membrane components compared to normal A31 cells. These differences consisted of increased amounts of simpler gangliosides, absence of the large external transformation sensitive (LETS) protein, and the appearance of a major new glycoprotein band of about 105 000 molecular weight. In contrast, the spontaneously transformed cell line that caused the fastest growing tumors in vivo and the most rapid animal death (3T12T) did not have these changes. A31 and 3T12T glycolipid profiles appear similar as did glycoproteins and cell surface proteins detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. When Pronase-generated glycopeptides were analyzed by Sephadex G-50 chromatography, and enrichment in faster-eluting species was seen in two killing tumor lines (c5T and 3T12T) compared to A31. Regressing tumor lines (MSC, c5) did not show this change. Isolated membrane glycoproteins yield glycopeptides of different sized after Pronase digestion. In addition, several 3T12T glycoproteins yield glycopeptides that are larger than those from the corresponding glycoproteins of A31 cells. It appears that glycopeptide alterations associated with transformation occur in several membrane glycoproteins.

Pancreas-specific messenger RNAs, in particular that for amylase, have been measured during the course of development. In addition the modulation of specific gene expression during differentiation in vitro by bromodeoxyuridine and the glucocorticoid, dexamethasone, has been examined. BrdU depresses the accumulation of all of the tissue-specific mRNAs. Dexamethasone selectively increases the concentration of amylase mRNA. Changes in the levels of mRNAs correlate directly with changes in the levels of pancreas-specific proteins. Pancreatic rudiments release into the incubation medium a characteristic subset of cellular proteins. Some of these, including amylase, represent the secretory digestive (pro)enzymes. Another group of proteins are also released by immature rudiments which have not yet accumulated the digestive enzymes. Some of these proteins persist throughout development and may be diagnostic of and functionally associated with pancreatic differentiation.