Mouse Peritoneal Fluid Research Articles

The cytochrome P4501A1 (CYP1A1) inhibitor bergamottin enhances host tolerance to multidrug-resistant Vibrio vulnificus infection

PurposeVibrio vulnificus (V. Vulnificus) infection is characterized by rapid onset, aggressive progression, and challenging treatment. Bacterial resistance poses a significant challenge for clinical anti-infection treatment and is thus the subject of research. Enhancing host infection tolerance represents a novel infection prevention strategy to improve patient survival. Our team initially identified cytochrome P4501A1 (CYP1A1) as an important target owing to its negative modulation of the body's infection tolerance. This study explored the superior effects of the CYP1A1 inhibitor bergamottin compared to antibiotic combination therapy on the survival of mice infected with multidrug-resistant V. Vulnificus and the protection of their vital organs. MethodsAn increasing concentration gradient method was used to induce multidrug-resistant V. Vulnificus development. We established a lethal infection model in C57BL/6J male mice and evaluated the effect of bergamottin on mouse survival. A mild infection model was established in C57BL/6J male mice, and the serum levels of creatinine, urea nitrogen, aspartate aminotransferase, and alanine aminotransferase were determined using enzyme-linked immunosorbent assay to evaluate the effect of bergamottin on liver and kidney function. The morphological changes induced in the presence of bergamottin in mouse organs were evaluated by hematoxylin and eosin staining of liver and kidney tissues. The bacterial growth curve and organ load determination were used to evaluate whether bergamottin has a direct antibacterial effect on multidrug-resistant V. Vulnificus. Quantification of inflammatory factors in serum by enzyme-linked immunosorbent assay and the expression levels of inflammatory factors in liver and kidney tissues by real-time quantitative polymerase chain reaction were performed to evaluate the effect of bergamottin on inflammatory factor levels. Western blot analysis of IκBα, phosphorylated IκBα, p65, and phosphorylated p65 protein expression in liver and kidney tissues and in human hepatocellular carcinomas-2 and human kidney-2 cell lines was used to evaluate the effect of bergamottin on the nuclear factor kappa-B signaling pathway. One-way ANOVA and Kaplan-Meier analysis were used for statistical analysis. ResultsIn mice infected with multidrug-resistant V. Vulnificus, bergamottin prolonged survival (p = 0.014), reduced the serum creatinine (p = 0.002), urea nitrogen (p = 0.030), aspartate aminotransferase (p = 0.029), and alanine aminotransferase (p = 0.003) levels, and protected the cellular morphology of liver and kidney tissues. Bergamottin inhibited interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α expression in serum (IL-1β: p = 0.010, IL-6: p = 0.029, TNF-α: p = 0.025) and inhibited the protein expression of the inflammatory factors IL-1β, IL-6, TNF-α in liver (IL-1β: p = 0.010, IL-6: p = 0.011, TNF-α: p = 0.037) and kidney (IL-1β: p = 0.016, IL-6: p = 0.011, TNF-α: p = 0.008) tissues. Bergamottin did not affect the proliferation of multidrug-resistant V. Vulnificus or the bacterial load in the mouse peritoneal lavage fluid (p = 0.225), liver (p = 0.186), or kidney (p = 0.637). ConclusionBergamottin enhances the tolerance of mice to multidrug-resistant V. Vulnificus infection. This study can serve as a reference and guide the development of novel clinical treatment strategies for V. Vulnificus.

Transcriptomics Changes in the Peritoneum of Mice with Lipopolysaccharide-Induced Peritonitis.

Peritonitis caused by LPS is a severe clinical challenge, which causes organ damage and death. However, the mechanism of LPS-induced peritonitis has not been fully revealed yet. Here, we investigated the transcriptome profile of the peritoneal tissue of LPS-induced peritonitis in mice. A model of LPS-induced peritonitis in mice was established (LPS 10 mg/kg, i.p.), and the influence of TAK 242 (TLR4 inhibitor) on the level of inflammatory cytokines in mouse peritoneal lavage fluid was investigated by using an ELISA test. Next, the peritoneal tissues of the three groups of mice (Control, LPS, and LPS+TAK 242) (n = 6) were isolated and subjected to RNA-seq, followed by a series of bioinformatics analyses, including differentially expressed genes (DEGs), enrichment pathway, protein-protein interaction, and transcription factor pathway. Then, qPCR verified-hub genes that may interact with TAK 242 were obtained. Subsequently, the three-dimensional structure of hub proteins was obtained by using homology modeling and molecular dynamics optimization (300 ns). Finally, the virtual docking between TAK 242 and hub proteins was analyzed. Our results showed that TAK 242 significantly inhibited the production of inflammatory cytokines in the peritoneal lavage fluid of mice with peritonitis, including IL-6, IFN-γ, IL-1β, NO, and TNF-α. Compared with the Control group, LPS treatment induced 4201 DEGs (2442 down-regulated DEGs and 1759 up-regulated DEGs). Compared with the LPS group, 30 DEGs were affected by TAK 242 (8 down-regulated DEGs and 22 up-regulated DEGs). A total of 10 TAK 242-triggered hub genes were obtained, and the possible docking modes between TAK 242 and hub proteins were acquired. Overall, our data demonstrated that a large number of DEGs were affected in LPS-triggered peritonitis mice. Moreover, the TLR4 inhibitor TAK 242 is capable of suppressing the inflammatory response of LPS-induced peritonitis. Our work provides clues for understanding the pathogenesis of LPS-induced peritonitis in mice.

CpG-DNA exerts antibacterial effects by protecting immune cells and producing bacteria-reactive antibodies

CpG-DNA activates various immune cells, contributing to the host defense against bacteria. Here, we examined the biological function of CpG-DNA in the production of bacteria-reactive antibodies. The administration of CpG-DNA increased survival in mice following infection with methicillin-resistant S. aureus and protected immune cell populations in the peritoneal cavity, bone marrow, and spleen. CpG-DNA injection likewise increased bacteria-reactive antibodies in the mouse peritoneal fluid and serum, which was dependent on TLR9. B cells isolated from the peritoneal cavity produced bacteria-reactive antibodies in vitro following CpG-DNA administration that enhanced the phagocytic activity of the peritoneal cells. The bacteria-reactive monoclonal antibody enhanced phagocytosis in vitro and protected mice after S. aureus infection. Therefore, we suggest that CpG-DNA enhances the antibacterial activity of the immune system by protecting immune cells and triggering the production of bacteria-reactive antibodies. Consequently, we believe that monoclonal antibodies could aid in the treatment of antibiotic-resistant bacterial infections.

Quorum-sensing molecules N-acyl homoserine lactones inhibit Trueperella pyogenes infection in mouse model

Trueperella pyogenes is a gram-positive opportunistic pathogen normally causes mastitis, liver abscesses and pneumonia of economically important livestock. It has been suggested that gram-negative bacteria can suppress the growth and virulence of T. pyogenes in vitro by using the quorum-sensing (QS) signal molecules and cause the transition of predominant species. However, whether these QS signals can be used as potential anti-virulence drugs against T. pyogenes infection is unclear. In this study, the in vivo inhibitory effect N-acyl homoserine lactones (AHLs) from Escherichia coli and Pseudomonas aeruginosa on T. pyogenes was tested by using mouse model. Mice were first peritoneally infected with T. pyogenes followed by intravenous injection of N-Octanoyl-DL-homoserine lactone (C8HSL) or N-(3-oxododecanoyl) homoserine-l-lactone (C12HSL). The results showed that C8HSL and C12HSL significantly reduced bacterial load and increased the survival rate of mice against T. pyogenes challenge. Additionally, the treatment of AHLs promoted the secretion of IL-1β, IL-6, IL-8 and TNF-α in mouse peritoneal fluid, and significantly decreased the expression levels of virulence genes of residual T. pyogenes. Importantly, murine macrophages rapidly phagocytosed bacteria when they were treated with AHLs compared to untreated cells. Collectively, our findings provide a major advance in understanding the inhibitory effect of AHLs in vivo and a promise for developing new clinical or veterinary treatments of T. pyogenes-related infection.

Acute phase inflammation is characterized by rapid changes in plasma/peritoneal fluid N-glycosylation in mice.

Murine zymosan-induced peritonitis is a widely used model for studying the molecular and cellular events responsible for the initiation, persistence and/or resolution of inflammation. Among these events, it is becoming increasingly evident that changes in glycosylation of proteins, especially in the plasma and at the site of inflammation, play an important role in the inflammatory response. Using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS)-based glycosylation profiling, we investigated the qualitative and quantitative effect of zymosan-induced peritonitis on N-glycosylation in mouse plasma and peritoneal fluid. Our results show that both N-glycomes exhibit highly similar glycosylation patterns, consisting mainly of diantennary and triantennary complex type N-glycans with high levels (>95 %) of galactosylation and sialylation (mostly NeuGc) and a medium degree of core fucosylation (30 %). Moreover, MS/MS structural analysis, assisted by linkage-specific derivatization of sialic acids, revealed the presence of O-acetylated sialic acids as well as disialylated antennae (“branching sialylation”) characterized by the presence of α2-6-linked NeuGc on the GlcNAc of the NeuGcα2-3-Galβ1-3-GlcNAc terminal motif. A significant decrease of (core) fucosylation together with an increase of both α2-3-linked NeuGc and “branching sialylation” were observed in N-glycomes of mice challenged with zymosan, but not in control mice injected with PBS. Importantly, substantial changes in glycosylation were already observed 12 h after induction of peritonitis, thereby demonstrating an unexpected velocity of the biological mechanisms involved.Electronic supplementary materialThe online version of this article (doi:10.1007/s10719-015-9648-9) contains supplementary material, which is available to authorized users.

Isolation of Mouse Neutrophils.

Neutrophils represent the first line of defense against bacterial and fungal pathogens. Indeed, patients with inherited and acquired qualitative and quantitative neutrophil defects are at high risk for developing bacterial and fungal infections and suffering adverse outcomes from these infections. Therefore, research aiming at defining the molecular factors that modulate neutrophil effector function under homeostatic conditions and during infection is essential for devising strategies to augment neutrophil function and improve the outcome of infected individuals. This unit describes a reproducible density gradient centrifugation-based protocol that can be applied in any laboratory to harvest large numbers of highly enriched and highly viable neutrophils from the bone marrow of mice both at the steady state and following infection with Candida albicans as described in UNIT. In another protocol, we also present a method that combines gentle enzymatic tissue digestion with a positive immunomagnetic selection technique or Fluorescence-activated cell sorting (FACS) to harvest highly pure and highly viable preparations of neutrophils directly from mouse tissues such as the kidney, the liver or the spleen. Finally, methods for isolating neutrophils from mouse peritoneal fluid and peripheral blood are included. Mouse neutrophils isolated by these protocols can be used for examining several aspects of cellular function ex vivo including pathogen binding, phagocytosis and killing, neutrophil chemotaxis, oxidative burst, degranulation and cytokine production, and for performing neutrophil adoptive transfer experiments.

Deficiency of Immunophilin FKBP52 Promotes Endometriosis

Endometriosis is a common gynecological disease that affects approximately 10% of women of childbearing age. It is characterized by endometrial growth outside the uterus and often results in inflamed lesions, pain, and reduced fertility. Although heightened estrogenic activity and/or reduced progesterone responsiveness are considered to be involved in the etiology of endometriosis, neither the extent of their participation nor the underlying mechanisms are clearly understood. Heterogeneous uterine cell types differentially respond to estrogen and progesterone (P(4)). P(4), primarily acting via its nuclear receptor (PR), activates gene transcription and impacts many reproductive processes. Deletion of Fkbp52, an immunophilin cochaperone for PR, results in uterine-specific P(4) resistance in mice, creating an opportunity to study the unique aspects of P(4) signaling in endometriosis. Here we explored the roles of FKBP52 in this disease using Fkbp52(-/-) mice. We found that the loss of FKBP52 encourages the growth of endometriotic lesions with increased inflammation, cell proliferation, and angiogenesis. We also found remarkable down-regulation of FKBP52 in cases of human endometriosis. Our results provide the first evidence corroborated by genetic studies in mice for a potential role of an immunophilin cochaperone in the etiology of human endometriosis. This investigation is highly relevant for clinical application, particularly because P(4) resistance is favorably indicated in endometriosis and other gynecological diseases.

Toxoplasma gondii: In vivo and In vitro Cystogenesis of the Virulent RH Strain

Models of cystogenesis were developed in vivo and in vitro with the virulent RH Toxoplasma gondii strain. In vivo in a first assay, 7.5-wk-old Fischer rats were inoculated intraperitoneally with 10(7) tachyzoites from mouse peritoneal fluid. Between 9 and 17 wk after inoculation, 1-5 cysts of 40-60 microns, containing numerous bradyzoites, were isolated from the brain of 5 of the rats. In a second assay, 24 Wistar rats were inoculated intraperitoneally with 10(2), 10(4), and 10(8) tachyzoites at 7, 11, 24, and 46 days. An inoculum of 10(8) was lethal for the 6 rats inoculated before 46 days and there was no correlation between the inoculum and the number of cysts except in the rats inoculated at 46 days. Fibroblast cell cultures (MRC5), at pH 7.5, were destroyed in 72 hr by 10(5)/ml RH-strain tachyzoites. However, when pH culture was increased to pH 8, up to 140 cysts were observed with indirect fluorescent antibody staining. Electron microscopy showed a 15 microns cyst, containing bradyzoites and surrounded by a 0.5-1-micron irregular wall.

Antigen analysis of Toxoplasma gondii lysate and excretory-secretory materials by enzyme-linked immunoelectrotransfer blot (EITB)

Recently, the importance of toxoplasmosis is raised as a complication in immunosuppressed or AIDS patients. Our study focused on the identification of a variety of Toxoplasma antigens by immunoblotting. Rabbits and BALB/c mice were immunized with Toxoplasma lysate (RH strain), frozen tachyzoites (RH strain) or cysts (Beverly and Fukaya strain). Blood were collected from ear vein, heart or orbital plexus for detecting the serum antibody levels. For excretory-secretory (E.S) antigens, T. gondii (RH) tachyzoite were cultured in CHL (Chinese hamster lung) cells with MEM containing of 5% FCS. After 72 hrs, culture supernatant was collected. BALB/c mice were inoculated with RH tachyzoite intraperitoneally and peritoneal fluids were extracted three days later. E.S antigens were detected in culture supernatant and infected mouse peritoneal fluid by EITB. Serum IgG levels in rabbit were 1:512 of 10 days after primary immunization, 1:2,048 of 10 days after secondary immunization, 1:1,024 of 20 days after secondary immunization by IFAT, respectively. Serum IgG levels of immunized mice were 1:128 after 7 weeks. Tachyzoite antigens of the RH strain were detected 25 protein bands ranging 10 kDa-220 kDa of molecular weights with Coomassie blue stain. Toxoplasma major antigens corresponding to MW of 24 kDa, 27 kDa, 30 kDa, 35 kDa, 38 kDa were recognized by IgG and IgM antibodies. Excretory-secretory antigens present in culture supernatant with M. W. of 20, 30 kDa and in infected mouse peritoneal fluid with M.W. of 33 (P30), 45 kDa. When RH tachyzoite antigen was probed with different mice sera immunized with 2 strains of T. gondii, the IgG antibody band of Fukaya and Beverly strain (8 week-serum) is identical to those of RH strain. It is considered that the 30 kDa polypeptide detected in excretory-secretory materials and lysate was important major antigen of T. gondii (RH).

In vitro Damaging Effect of Leukocytes, Serum, and Peritoneal Fluid on Ascaris suum Infective Larvae

Peritoneal leukocytes, serum, and peritoneal fluid from Ascaris suum-infected or noninfected mice were studied using 51Cr release and larval mobility to determine if they had in vitro cytotoxicity for A. suum infective larvae. Mouse serum or peritoneal fluid from infected and non-infected mice was added to leukocyte-larvae cultures to determine their effect on leukocyte damage to larvae. Leukocytes from A. suum infected mice were cytotoxic for A. suum larvae in the presence of serum from infected or noninfected mice but only serum from infected mice produced larval cytotoxicity independent of leukocytes or in the presence of leukocytes from noninfected mice. Mouse peritoneal fluid from infected or noninfected mice was found to damage larvae independent of leucocytes. Cytotoxic activity was blocked when serum or peritoneal fluid was heat-inactivated suggesting complement as the damaging agent.

Effect of Extracellular Ions on Motility and Cell Entry in Toxoplasma gondii

Toxoplasma gondii tachyzoites were quiescent in mouse peritoneal fluid or in K2SO4 buffer at pH 8.2. They became consistently motile when K+ was replaced by other monovalent or divalent cations at a constant pH (pH = 8.2). They also became motile when Cl- was substituted for SO4(2-). Nitrate or SCN-, can also be substituted for Cl- to a certain extent. Tachyzoites showed independent movement for more than 15 min in KCl, and for about 5 min in the other buffers at pH 8.2 after which they were exhausted and stopped. These tachyzoites could not then be further stimulated to motility by renewal of the suspension buffer. Infection of monolayer cells was demonstrated only with parasites which were motile during inoculation. The highest infectivity was thus obtained either with freshly collected tachyzoites or with those preincubated in K2SO4 buffer for 30 min at 37 degrees C at alkaline pH and thus not yet exhausted for motility. Approximately 34 to 38% of these latter organisms were seen to enter cells when they were inoculated into cultures immediately after being resuspended in MEM for 30 min at 37 degrees C. Conversely, those whose motility had been exhausted by the preincubation in buffers other than K2SO4, pH 8.2 could not enter monolayer cells. Additionally, parasites were unable to enter cells when inoculated into cultures in K2SO4 buffer at alkaline pH; instead they remained quiescent on the surface of the monolayer cells, suggesting that Toxoplasma enters the host cells by active invasion.

Effects of extracellular potassium on acid release and motility initiation in Toxoplasma gondii.

The internal pH (pHi) of Toxoplasma gondii was estimated by measuring the accumulation of the weak base 9-aminoacridine in buffers with various ionic compositions. The pHi of the metabolizing parasite increased when the extracellular K+ was elevated in alkaline medium or when the external pH (pHe) was substantially increased in medium employing high external K+ (90 mM). The parasite in mouse peritoneal fluid, or in potassium sulfate buffer (pH 8.2), where the pHi was demonstrated to be increased to 7.9, became motile when acidic buffer was substituted for the original suspension medium. This acid-induced independent movement subsided within 5 min but was repeatedly induced if the pHe was serially lowered to 6.0. Basic buffers, on the other hand, abolished motility when applied to the moving parasites. Nigericin, which is known to collapse pH gradients across the membrane, also abolished motility.

Fimbrial phase variation and systemic E. coli infection studied in the mouse peritonitis model

Mouse peritonitis induced by intraperitoneal injection of a virulent (LD 50 4 × 10 5) E. coli 018:K1:H7 strain isolated from neonatal meningitis was studied. These bacteria are capable of producing both type 1 and S fimbriae, binding to mannose or sialic acid containing glycoconjugates, respectively; the production of both fimbrial types is subject to phase variation. A broth culture of the bacteria was fractionated into subpopulations containing either type 1 or S fimbriae or neither (nonfimbriated cells), and each fraction, grown in broth to logarithmic growth phase, was used to infect groups of mice. The type 1 fraction was associated with decreased virulence as the fraction was eliminated rapidly without causing a progressive infection even at 10 6 bacteria/mouse, whereas both S and nonfimbriated cells started rapid multiplication in the peritoneal cavity and spread to the blood. In nonfimbriated cells, however, S fimbriae production was induced at the same time so that at 1 h after injection, 60–70% of the bacteria in the peritoneal cavity and in the blood of the mice had S fimbriae. The injected S-fimbriated fraction remained completely S-fimbriated. Rapid induction of S fimbriae also took place in vitro when the nonfimbriated bacteria were grown in mouse serum or peritoneal fluid. Anti-S serum protected the mice from a lethal dose of S-fimbriated bacteria.

Increased local complement levels upon intraperitoneal injection of mice with Listeria monocytogenes and regulation by polyanions

Alternative complement pathway activity in mouse peritoneal fluid was determined by a sensitive microtiter assay with rabbit erythrocytes as target cells and cobra venom factor as the inducer of bystander hemolysis. The intra- and interstrain variations in five mouse strains were 30% and, maximally, 64%, respectively. No activity was detected in genetically C5-deficient mice. In C5-sufficient mice, intraperitoneal injection of heat-inactivated Listeria monocytogenes resulted in a 250% increase in local complement activity within 30 min. This effect was reversed by simultaneous injection of dextran sulfate but not heparin. These results are discussed in relation to the ability of listeriae to activate the mouse alternative complement pathway in vitro and the effectiveness of dextran sulfate and the failure of heparin in functioning as an adjuvant in the induction of a protective response in mice by a heat-inactivated listeria vaccine.

Production of cytotoxic factor into mouse peritoneal fluid by OK-432, a streptococcal preparation

A cytotoxic factor was induced by the injection of LPS into the peritoneal fluids of mice which had been previously primed with a streptococcal antitumor preparation, OK-432. No cytotoxic effect on L-929 cells was observed in the peritoneal fluids of mice singly treated with OK-432 or LPS. Various mouse and human tumor cell lines were effectively killed by this peritoneal cytotoxic factor, though normal cell lines were insensitive, which indicates that this factor is not species-specific. The highest level of cytotoxic activity was obtained when LPS was given to mice 5 days after the injection of OK-432. The optimal time for collection of peritoneal fluids for the cytotoxic factor was 2 h following the LPS injection. Interferon activity was found to be negative by the plaque reduction test using L-929 cells with vesicular stomatitis virus. These results suggest that this cytotoxic factor is similar to the tumor necrosis factor (TNF) in the mouse serum.

ELEMENTAL ANALYSIS OF HUMAN PLATELETS ON FRESH AIR-DRIED BLOOD SMEARS BY WAVE DISPERSIVE X-RAY MICROANALYSIS USING STEM EQUIPMENT

Wave dispersive X-ray microanalysis was performed on platelets and red blood cells of a healthy male and a von Willebrand patient, and mast cells and lymphocytes of the mouse peritoneal fluid. A STEM equipment of target method was modified by replacing grid holders with carbon rod tubes and the reflecting stage with a carbon disc for microanalysis, attempting to decrease the background noises. In secondary electron images at acceleration voltage of 15-45kV and absorption current of 0.03-0.1nA, the above cells were recognized and also identified in back scattered electron images at an absorption current of 0.1-0.5μA by their profiles and relationship with other cells. High peaks of characteristic X-ray counts of Na-, P-, S-, Cl and K-Kα were disclosed in the analysis of cells. Ca- and Mg-Kα were distinctly disclosed in that of the healthy male and Ca- or Mg-Kα was not in that of patient platelets. Analysis of healthy male platelets, mouse mast cells and lymphocytes by means of the pulse height analyzers detected Zn-Kα and Zn-Lα. Electron microscopy of healthy male platelets disclosed the dense bodies but platelets of the patient failed to reveal them.

Regulation of bone marrow cell growth in diffusion chambers: the effect of adding normal and leukemic (CML) polymorphonuclear granulocytes

Inhibition of granulopoiesis was studied using the diffusion chamber (DC) technique. When mature granulocytes from human blood or syngeneic mouse peritoneal fluid were added to mouse bone marrow cells cultured in DC, a significant depression of granulopoiesis took place, and a stimulation of macrophage formation was observed in 7-day cultures. Human granulocytes had a stronger inhinitory capacity than mouse granulocytes. The inhibition appeared to be tissue-specific and caused by a diffusible factor. A time study showed that the added granulocytes had no observable effect on the growth of proliferative granulocytes and CFU-C during the first days of culture. A rapid decrease of proliferative granulocytes after day 5 was preceded by a similar reduction of CFU-C 1 day earlier. The effect of CFU-S was more variable. In one strain of mice, there was a consistent increase in the granulocyte co-culture group, whereas in another strain a significant increase was observed only on day 2. Hislotologic examination showed that mature granulocytes changed the colony distribution, so that a significant relative decrease of granuloid colonies occurred. The nature of this delayed suppression of granulopoiesis is not evident from these data. A possible explanation is that factors released by mature granulocytes prevent recruitment of CFU-C and granulocyte precursors from the CFU-S compartment by blocking the granulopoietic pathway. Leukemic (CML) granulocytes isolated from blood were less able to inhibit granulopoiesis than normal granulocytes with mouse bone marrow as well as human bone marrow as target cells.

Regulation of Bone Marrow Cell Growth in Diffusion Chambers: The Effect of Adding Normal and Leukemic (CML) Polymorphonuclear Granulocytes

Inhibition of granulopoiesis was studied using the diffusion chamber (DC) technique. When mature granulocytes from human blood or syngeneic mouse peritoneal fluid were added to mouse bone marrow cells cultured in DC, a significant depression of granulopoiesis took place, and a stimulation of macrophage formation was observed in 7-day cultures. Human granulocytes had a stronger inhinitory capacity than mouse granulocytes. The inhibition appeared to be tissue-specific and caused by a diffusible factor. A time study showed that the added granulocytes had no observable effect on the growth of proliferative granulocytes and CFU-C during the first days of culture. A rapid decrease of proliferative granulocytes after day 5 was preceded by a similar reduction of CFU-C 1 day earlier. The effect of CFU-S was more variable. In one strain of mice, there was a consistent increase in the granulocyte co-culture group, whereas in another strain a significant increase was observed only on day 2. Hislotologic examination showed that mature granulocytes changed the colony distribution, so that a significant relative decrease of granuloid colonies occurred. The nature of this delayed suppression of granulopoiesis is not evident from these data. A possible explanation is that factors released by mature granulocytes prevent recruitment of CFU-C and granulocyte precursors from the CFU-S compartment by blocking the granulopoietic pathway. Leukemic (CML) granulocytes isolated from blood were less able to inhibit granulopoiesis than normal granulocytes with mouse bone marrow as well as human bone marrow as target cells.

Changes in cellular patterns of mouse peritoneal fluid with dextran administration.

Changes in cellular patterns of mouse peritoneal fluid with dextran administration.

Effect of sodium chloride on female mouse peritoneal fluid cell content.

Une heure apres injection i.p. d'une solution de 1% de chlorure de sodium, nous avons observe un grand changement dans le pourcentage des diverses cellules du liquide peritoneal de souris femelles adultes. Nous envisageons ainsi la possibilite d'irriguer la cavite abdominale avec une solution saline tant en vue d'etudes cliniques qu'experimentales pour obtenir le liquide en question.