The phenotype of alveolar-associated fibroblasts (Afb) in sarcoidosis (SA) and idiopathic pulmonary fibrosis (IPF) is unclear. In the present study, we characterized the cytoskeletal proteins and the contraction properties in alveolar-associated fibroblasts recovered by bronchoalveolar lavage (BAL) in the two diseases. Afb were studied from BAL cells in eight IPF and seven SA patients. Cytoskeletal proteins were identified by ELISA and immunofluorescent methods. Biochemical measurements were done by dry chemistry. Contraction was performed by a gel contraction assay. Afb alpha-SM actin measured by ELISA was higher in IPF than in SA (p = 0.042). Vimentin, desmin, myosin, and fibroblast markers were expressed equally. Only in IPF did the Afb reveal the myofibroblast phenotype showing alpha-SM actin immunofluorescence labeling and, by electron microscopy, filaments with associated dense bodies with rough endoplasmic reticulum. Gel contraction showed that cells in IPF contracted significantly more than in SA (p = 0.046 IPF versus SA). The addition of ET-1 increased contraction in all groups. Dry chemistry analysis showed higher levels (p = 0.0065) of creatine phosphokinase (CPK), lower levels of glucose (p = 0.0082), and similar levels of Ca(2+) and lactate in the IPF and SA Afb. Dinitrofluorobenzene (DNFB), a potent inhibitor of CPK, completely abolished spontaneous cell contraction. Afb differentiates into myofibroblasts with different biochemical and energetic properties in IPF. Moreover, Afb from IPF patients showed increased contractile properties. This may explain the difference in the behavior patterns and outcomes of the two diseases.

The anticancer chemotherapeutic drugs melphalan (L-phenylalanine mustard; L-PAM), 5-fluorouracil (5-FU), methotrexate (MTX), and daunorubicin (DAU) were tested for their toxic activity against MOPC-315 tumor cells in vitro. L-PAM, 5-FU, and DAU had a marked toxic effect whereas MTX did not affect the rate of thymidine incorporation in the tumor cells. L-PAM (7.5 mg/kg) induced permanent regression of large s.c. MOPC-315 plasmacytoma tumors, 5-FU (200-250 mg/kg) induced transient regression of MOPC-315 tumors with reappearance starting on the 6th day after the 5-FU injection and DAU (5 mg/kg) was not effective. L-PAM treatment restored the cytotoxic potential of spleen cells of tumor-bearing mice against target MOPC-315 tumor cells whereas spleen cells from tumor-bearing mice treated with 5-FU were unable to mount a cytotoxic response. L-PAM and 5-FU were also assayed for their effect in vitro on induction of suppressor T cells by ConA. L-PAM treatment in vitro markedly reduced the induction of suppressor T cells by ConA whereas 5-FU had no effect. It is suggested that anticancer chemotherapeutic drugs can be classified in "immunopromoting" (L-PAM as prototype) and "nonimmunopromoting" (5-FU as prototype) on the basis of their effect in vivo on established tumors and their effect on induction of suppressor T cells by ConA.

Mice cured from large MOPC-315 tumors by a single dose of melphalan, 7.5 mg/kg, were examined for up to 60 days after the drug treatment (71 days after the tumor inoculation) for their ability to respond to mitogenic stimulation, specific and nonspecific antigenic stimulation and for their susceptibility to inoculation with an unrelated tumor, L10 lymphoma. The response of spleen cells from cured mice to mitogenic stimulation by phytohemagglutinin or concanavalin A was slightly depressed at an early stage after the drug treatment. The allogeneic response against C57BL spleen cells and the antibody response against sheep red blood cells (SRBC) of spleen cells from cured mice remained below normal levels during the whole observation period. The deficiency in response to antigenic stimulation was found to be due to impairment in T-cell function. Cured mice were also deficient in their response to SRBC immunization (antibody and delayed-type hypersensitivity responses) and were more susceptible to inoculation with an unrelated tumor, L10 lymphoma, than normal, noninoculated mice. On the other hand, spleen cells of cured mice developed a highly specific cytotoxic response against target MOPC-315 tumor cells and the cured mice were resistant to challenge with an otherwise highly tumorigenic dose of MOPC-315. Thus, cured mice remained deficient for a long period of time in their response to MOPC-315-unrelated antigens but, at the same time, they showed a potent specific antitumor immunity potential in vivo and in vitro.

In the present study we aimed to detect early intracellular changes in the cytoplasmic matrix induced in human, pulmonary-derived fibroblasts following exposure to interleukin (IL)-1α IL-1β and tumor necrosis factor-α. Such changes were detected by measuring intracellular fluorescein fluorescence polarization (IFFP) using the Cellscan apparatus. IFFP measurement was selected in our study since it has been shown to reflect the microviscosity of the cytoplasmic matrix. Significant reductions ( > 5%) in the IFFP were induced in fibroblasts by all the cytokines employed. The effect of cytokines on IFFP was achieved at concentrations of 5-10 ng/ml of the cytokines. The reduction in IFFP, following stimulation with the cytokines, was detected as early as 20 min after exposure to the cytokines, lasted at least 40-60 min after exposure to IL-1α and IL-1β, and was inhibited by vinblastine, an inhibitor of the polymerization of microtubules. Our results show that IFFP measurements by the Cellscan may reveal rapid intracellular changes occurring in the cytoskeleton components of activated cells.