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The effect of Peptichemio (PTC) on the survival and DNA synthesis in mouse L-cells was studied. Experimental data show that a typical dose-survival curve is obtained when L-cells are cultivated in the presence of PTC. Exposure of L-cells to various concentrations of PTC caused an inhibition in cellular ability to synthesize DNA which is evidenced by the decreased incorporation of 3H-thymidine. This inhibition is dependent on the concentration of PTC and the time of exposure.

The PHA-responsiveness in cell-culture of mononuclear cells obtained in acute leukemia from different sources, such as blood, bone marrow, lymph-nodes, leukemic pleural effusion and spinal fluid were investigated. The peripheral blood lymphocytes of all but 2 of 28 untreated patients showed extents of blastogenesis from 48 to 90%. The bone-marrow cells of 18 of the 28 patients gave high extents of blast-like cells (52 to 88%) in 5. In fact, the marrow cells of 9 patients showed blast extents only from 1 to 14%, whereas the remaining 4 patients had no marrow-cell transformation at all. Also the lymphocytes from lymph-nodes of 2 patients show low extents of blastogenesis (8.5–12.3%). Again, the mononuclear cells obtained from the spinal fluid in 7 patients affected by meningeal leukemia, and from the pleural effusion in 1 patient did not undergo any blastic transformation. The question arises whether the PHA-responsive cells from the acute leukemia peripheral blood are normal immuno-competent lymphocytes circulating in the blood, independent from the cell type of leukemia. As a consequence, it comes up whether the bone-marrow cells are PHA-responsive in the lymphocytic type of acute leukemia and PHA-nonresponsive in other cell-types of leukemia (myelo-monocytic). Thus, the PHA-cell culture of both the peripheral blood and the bone-marrow cells could aid in recognizing the cell types of leukemia one is dealing with.

Resume L'activite proliferative des plasmocytes et des plasmoblastes de la moelle de patients atteints de plasmocytome fut determinee par marquage a la thymidine tritiee in vitro. On observa une faible incorporation de l'isotope, indiquant une activite proliferative reduite des plasmocytes anormaux, similaire a celle qui a ete observee precedemment par l'etude des index mitotiques et stathmocinetiques. Cependant, les indices de proliferation etaient plus eleves dans les cellules les plus immatures, de sorte qu'il est necessaire que la maturite des cellules soit prise en consideration.

Histochemical observations on the iron content (hemosiderin) within sections of Brunner's glands in duodenal biopsies showed that practically no iron is contained within the glands of healthy persons. In contrast, in cases of marked transfusional hemosiderosis, the cuboidal cells of the same glands showed a high hemosiderin content. In the most severe cases, 100% of the tubules had a positive reaction to Perls's test. The intensity of the iron-positive reaction within the duodenal Brunner glands varied according to the severity of hemosiderosis. These results are in agreement with the interpretation that in hemosiderosis the desquamation of iron-loaded epithelial cells of Brunner's glands contributes to the removal of iron from the body. The cells have this function in common with the epithelium of other mucus-secreting glands and with macrophages.

Crosby's intestinal biopsies were performed on patients with acute epidemic hepatitis, both while the disease was in progress and after the patient had recovered. The mucosal biopsies were examined both by stereomicroscopy and by histologic section. While the disease was in progress, there were various degrees of villous atrophy, alterations of the covering epithelium, and cellular interstitial infiltration in the stroma and in the tunica propria. During recovery the findings were characterized by hypertrophied villi, thoroughly restored epithelium, and infiltration of mononuclear cells in the wall of the mucosa.

Bone marrow plasma cells from fifteen cases of multiple myeloma, immunologically typed, were incubated with different tritiated compounds. The labelling index with tritiated thymidine is generally low, while the mean grain count is fairly normal in the active cells. The labelling index of 3H-uridine and 3H-leucine was very high, while the mean grain count per cell lies within the normal range. The results obtained with 3H-phenylalanine-mustard (melphalan), which is a drug used in the treatment of the plasmacytoma, show also incorporation values roughly comparable to those of 3H-leucine. The present data seem to support the clinical use of melphalan as a compound that is actively incorporated into the plasma cells of plasmacytoma although inhibition of protein synthesis due to specific binding to protein was not demonstrated.

The observation that phytohaemagglutinin (P.H.A.) - blstogenesis of human peripheral blood lymphocytes is inhibited by « E. coli » - L-asparaginase (« Asn-ase ») suggested that this enzyme might cause cellular immunological depression. To obtain more information on the possible immunosuppressive properties of « Asn-ase » the effect of this agent on lymphocyte blastogenesis in mixed-lymphocyte cultures has now been investigated. This system provides a real histo-compatibility test. The study was carried out on blood from 24 subjects. Most were normal volunteers but a few were suffering from plasmocytoma (1), acute leukaemia (1) and chronic lymphatic leukaemia (2). Peripheral-blood lymphocytes were randomly mixed in pairs and cultured. One group of these cultures was taken as control and another group was added to « Asn-ase » to make a concentration of one I.U. per ml of culture medium. These cultures were incubated at 37°C for 7 days. The results were 18.33 ± 7.02 % immunologically activated lymphocytes in the control mixed cultures, as opposed to 1.00 ± 1.29 % in the cultures with « Asn-ase ». These results show that « Asn-ase » is capable of inhibiting the histoincompatibility reaction in a mixed-lymphocyte culture and they strongly support the interpretation that « Asn-ase » has cellular immunosuppressive properties.

Dannomycin is an antibiotic which interferes with the proliferative activity of somatic cells. The effect of Dannomycin on the proliferative activity of stem-cells developed from human blood lymphocytes in tissue culture with phytohemagglutinin was studied. The degree of proliferation was evaluated by the stathmokinetic test. Daunomycin caused an inhibitory effect, but the degree of inhibition was not proportional to the dose of the drug. Small doses, such as 1 jig/ml reduced the degree of proliferation by one third, whereas 3 fig/ml reduced that activity to one third in respect to the control.

Resume Les lymphocytes du sang peripherique de la leucemie lymphoide chronique (LLC) reagissent mal a la phytohemagglutinine en culture de tissu, tandis que les lymphocytes de la leucemie myeloide chronique reagissent comme les lymphocytes du sang normal. Il apparait que cette absence de reponse au stimulus P.H.A. des lymphocytes de la LLC ne depend pas directement des cellules memes, mais du plasma. En fait, le plasma de LLC ajoute a des lymphocytes de sang normal, en culture de tissu avec P.H.A., empeche leur transformation en cellules proliferantes semblables a des cellules blastiques. L'irradiation de la rate par rayons X peut supprimer cette action inhibitrice du plasma de la LLC.

Es wurde die immunologische Antwort von Lymphozyten aus dem peripheren Blut auf Phytohamagglutinin in Gewebekulturen bei chronischlymphatischer Leukamie vor und nach einer Milzbestrahlung untersucht. Vor der Bestrahlung trat, wie gewohnlich bei einer chronisch lymphatischen Leukamie ein sehr geringer Prozentsatz an Blasten auf. Nach der Milzbestrahlung konnten aber wesentlich hohere Prozentsatze von Blasten-ahnlichen Zellen festgestellt werden.

In 24 men aged 32 to 58 years with precancerous states of the larynx, i.e., leukoplakia, papillomas and pachydermia the peripheral blood lymphocytes were cytochemically stained for N-acetyl-beta-glucosaminidase, beta-glucuronidase, acid phosphatase and glycogen; and the neutrophils were stained for alkaline phosphatase, myeloperoxidase and lipids. The results were expressed in terms of the absolute counts of reaction-positive cells and of the activity index score. The serum immunoglobulins IgG, IgA and IgM were also determined by Mancini's method. The results obtained were compared with those in 20 healthy men aged 20 to 30 years. The patients exhibited elevated numbers of N-acetyl-beta-glucosaminidase and beta-glucuronidase-positive lymphocytes. A characteristic feature was an increase in the absolute counts of lymphocytes with diffuse and granular-diffuse types of cytochemical reaction for all enzymes studied. The number of cells with the granular type of enzymatic reaction (intact enzyme-positive lysosomes) was significantly diminished. These cytochemical alterations were accompanied by a significant increase in the serum IgA level. These results are discussed with reference to the lymphoid system response to tissues of precancerous lesions of the larynx. So far as the neutrophils are concerned the patients exhibited significant intracellular deficiency of beta-glucuronidase and decrease in the lipid content as well as an elevated alkaline phosphatase activity. The possible significance of the beta-glucuronidase deficiency in neutrophils for the diminished cytotoxic response of these cells against the tumor and precancerous lesion cells is discussed.

In 20 men, age 35 to 55 years, with cancer of the larynx, activity of N-acetyl-beta-glucosaminidase in peripheral blood lymphocytes was determined cytochemically according to Hayashi's method. Numbers of enzyme-positive cells were counted with regard to the type of cytochemical reaction (granular, granular-diffuse, or diffuse), and to the number of enzyme-positive lysosomal granules within a single cell. In comparison to 20 healthy men, age 20 to 30 years, the following changes were noted in the patients: 1) the decrease of the total count of enzyme-negative lymphocytes; 2) the decrease of the total count of lymphocytes with granular type of reaction; 3) the increase of the counts of lymphocytes with granular-diffuse and diffuse type of reaction. The authors suggest that these changes could be related to the immune reaction of specific tumor antigens.

Using the cytochemical method after Barka and Anderson, activity and localization of lysosomal acid phosphatase (AP) was determined in peripheral-blood lymphocytes from 20 healthy subjects, 10 patients with Hodgkin's disease (HD), 10 patients with plasma cell myeloma (PCM), and 10 patients with primary polycythemia (PP). Total count of AP-positive lymphocytes was lowest in the patients with HD. Moreover, they showed a significant decrease of the absolute count of lymphocytes with well formed and more numerous AP-positive lysosomal granules. Analogous alterations in lymphocytes from patiens with PCM and those with PP were much less significant. The authors discuss the importance of the above-mentioned observations for evaluation on a cellular basis of lowered immunity of patients with lympho-proliferative and myelo-proliferative disorders, with special regard to AP as a T-cell marker.

The concept that the lymphoid tissue of the gastrointestinal tract is primarily involved in the immune defense mechanism has been repeatedly maintained by several investigators [1, 2]. Activation of intestinal lymphocytes to large pyroninophilic cells has been observed in cases where the lymphocytes, after crossing the basement membrane, have penetrated between or into the cells of the covering epithelium of intestinal villi [3, 4, 5, 6]. Such lymphatic transformation has been interpreted as an immunological reaction, but the most overwhelming evidence of the immunological function of the intestinal lymphoid tissue was provided by studies of Good and co-workers [7, 8, 9, 10], showing that the Peyer’s-patch-type of intestinal lymphoid tissue represents the mammalian counterpart of the bursa Fabricii in birds. On the other hand, it has been shown that the plasma cells of the intestinal mucosa constitute the major source of gamma-A immunoglobulins [11] and that gamma-A and gamma-M immunoglobulins are significantly increased in infectious hepatitis [12], which has been considered a generalized disease [13, 14]. Because the jejunal mucosa in infectious hepatitis occasionally shows a partial villous atrophy and frequently shows a marked cellular infiltration within the tunica propria [15], we undertook the present investigation. This study was performed to ascertain whether the intestinal infiltration contains immunologically activated cells, to determine the quantity of these cells, and to investigate the behavior of the lymphatic follicles and germinal centers of the duodenojejunal mucosa during the course of infectious hepatitis. These findings could be related to recent concepts of the immunological function of the lymphoid tissue in gastrointestinal tract.

The frequency, distribution, and morphologic characteristics of the mast cell were studied in jejunal biopsy specimens from normal subjects. A simple method of quantifying cells in intestinal specimens was described. Mast cells were found in variable numbers in the submucosa while in the lamina propria they may be present in large numbers or be totally absent. The different morphologic characteristics of mast cells in each of these areas was described and was attributed to a different functional state of activity of the cell. A significant decrease in the number of mast cells was observed in jejunal specimens of patients recovering from infectious hepatitis.

During the few last years the immune reaction of the germinal center to an antigen and/or immuno-depressive drug was extensively studied. The present experiment was designed to investigate the immune reaction of the germinal center induced by the enzymatic drug L-asparaginase. This compound can: 1st, depress lymphocyte blastogenesis (1), and, consequently, all the reactions connected with cell-immunity (2). This depression of lymphocyte blastogenesis is caused by quantities of L-asparaginase sufficient to deplete the exogenous L-asparagine, which is the specific substrate for L-asparaginase. 2nd, depress the antibody production, i.e., the humoral-immunity (3–5). Since the inhibition of antibody production occurs only after administration of large doses of L-asparaginase, this effect is probably not mediated by specific L-asparaginase activity. On the other hand, the administration of intermediate doses of that enzymatic compound induces the formation of specific anti-asparaginase antibodies (6,7).

The present experiments show that antimitotic agents commonly used in the treatment of neoplastic disorders do not behave uniformly with respect to the action on the blastic transformation of lymphocytes. Furthermore, their action differs according to the experimental procedure, namely whether they are administeredin vivo or added directly to the biological systemin vitro. It can be observed however, that a close relationship exists in thein vitro experiment between the inhibition of the blastic transformation and the antiproliferative effect, as measured by the inhibition of the uptake of tritiated thymidine. It is legitimate to believe that such agents are endowed with different degrees of immunological activity, which may be restrained or elicited according to the circumstances. Such effect should therefore be kept in mind, whenever these agents are given for a clinical treatment.

From observations, we conclude that in hepatic cirrhosis the duodenal and jejunal mucosa demonstrate the following profound alterations: (1) intense stasis, especially in the axis of the villus; (2) edema of the villus, with degeneration of the basement membrane and separation of the epithelium from the stroma; (3) degeneration of epithelium and stroma; (4) interstitial inflammation (small round-cell infiltration); (5) fibrous replacement of the stroma; (6) destruction of the walls of the vessels; and (7) dilatation of the crypts of Galeazzi-Lieberkuhn, with degeneration and atrophy of the cryptic epithelium and fibrosis. It is conceivable that these alterations taken together follow one another in the sequence described. The stasis of the intestinal mucosa might indeed result from portal stasis. This could be the fundamental cause of the alterations. Although this interpretation seems reasonable to us we cannot completely subscribe to it, because we have not excluded the possibility that the mucosa may have been damaged by the same factors that cause cirrhosis. Study of cases less advanced may clarify the mechanism of development of these lesions.

The effect of asparaginase on cellular immunity was investigated on human peripheral blood lymphocytes incubated in the culture system. Results showed that asparaginase inhibits lymphocyte blastogenesis, both when stimulated with PHA or PKW, and in the mixed lymphocyte cultures. This happened when the enzyme was added directly to the culture medium, as well as when the enzyme was administered i.v. in humans, and then the blood lymphocytes were removed and put into the PHA-culture medium added to the lymphocyte donor's plasma. Crossed experiments showed that: a) lymphocytes from asparaginase injected subjects which were set up in a PHA-culture-medium to which plasma (20%) from non-injected subjects had been added, undergo a normal amount of blastogenesis; and b) lymphocytes from non-injected subjects, set up in a PHA-culture-medium with plasma (20%) from asparaginase injected subjects show an inhibition in their blastic transformation.