Experimental Biology and Medicine

In the present minireview we have discussed several types of model systems which have given valuable clues to the ways in which leukemia and lymphoma cells mediate organ specific metastasis. Most patent is that these types of malignancies probably utilize normal receptor-mediated mechanisms for organ specific recognition. The animal models which have been most extensively studied indicate that the following three recognition mechanisms can be employed by metastatic lymphoma variants:

(1) Normal organ cells have receptors for determinants on metastatic variants 17

(2) metastatic variants have receptors for determinants expressed on endothelial cells 2 , 19 ; and

(3) an undefined interaction occurs between normal organ cells and metastatic variants 16 .

A great deal of evidence suggests that organ specific metastasis is at least partially mediated through carbohydrate determinants which are recognized by receptors. It is possible that these receptors may be part of a larger family of recognition molecules which mediate normal cell-cell interactions. Despite this, a role for a “microenvironmental” mechanism cannot be ruled out and, indeed, is not mutually exclusive of the receptor-mediated adherence mechanism. In fact, in our bone marrow model our preliminary results suggest the existence of growth factors which are secreted by adherent bone marrow cells and which strongly stimulate the growth of the leukemia cell lines. Nevertheless, the avid binding of leukemia cells to the adherent cells of the bone marrow appears to be an important mechanism whereby leukemia cells are “held” in the bone marrow. The continued development of MAbs which recognize organ specific receptors on tumor cells, the isolation of these receptors, and their molecular biology should reveal whether a multigene family for organ specific recognition exists. These receptors will serve as functional markers in contrast to conventional markers such as T- and B-cell determinants. While it is true that T- and B-cell markers may indicate the probable cell lineage of the transformant, they do not, for example, explain why ALL grows in the bone marrow and not in the spleen. We suggest that the identification of “homing” receptors using MAbs will, for the first time, enable us to gauge the status of a malignancy, accurately determine the prognosis, and even allow us to choose the appropriate MAb to be used for the therapy of individual leukemias.

Pneumococcal antibody responses and adverse reactions were assessed in 12 healthy volunteers who received either two (10 volunteers) or three (2 volunteers) doses of pneumococcal polysaccharide vaccines at 1 or 2 year intervals. The volunteers were given hexavalent (types 1, 3, 4, 7F, 8, 12F) octavalent (types 14, 19F added) nonavalent (type 5 added), and tridecavalent (types 6A, 6B, 9N, 18C, 23F added; type 5 removed) pneumococcal vaccines. Local and systemic reactions following any of the vaccines, either first, second, or third doses, were low, and these were mild; fever did not occur. Twelve volunteers received types 1, 3, 4, 7F, 8, and 12F in two doses of vaccine, and four volunteers received types 14 and 19F in two doses of vaccine. Primary immunization induced significant antibody rises to these antigens in most volunteers, but a second dose of vaccine did not further increase antibody levels to the same antigens; when it also contained new antigens, antibody rises usually developed to these antigens. Although the data are limited to detailed studies of only 12 persons, the results suggest that additional studies of this issue would seem appropriate.

The effect of various concentrations of acetaldehyde (0, 0.05, 0.1, 0.25, 0.5, 1.0, and 5.0 mM) on the relative rates of formation of hemoglobin acetaldehyde adducts detected in fractions eluted from cation exchange high-pressure liquid chromatography (HPLC) was investigated. When the hemoglobin and acetaldehyde mixtures were incubated at 37°C for various time intervals up to 24 hr, increased amounts of HbA1c could be observed after 2 hr incubation with 1 mM or greater concentrations of acetaldehyde, or after 4 hr incubation with at least 0.5 mM acetaldehyde. An increase in the HbA1a+b fraction was not observed with 4 hr incubation time until the acetaldehyde level reached 1 mM. The HPLC method detected no difference in minor hemoglobins from alcoholic and normal subjects. Incubation of red blood cells at 37°C for 1 hr with six consecutive pulses of 0.05 mM [¹⁴C]acetaldehyde showed no differences in the amounts of minor hemoglobins determined chromatographically at various pulse intervals. However, the measure of the ¹⁴C-label incorporation into hemoglobin showed that adducts eluting in the HbA1a+b fraction were formed at a faster rate than those eluting in the HbA1c or HbAo fraction, respectively. The specific activities of the HbA1a+b fractions at 2, 4, and 6 pulses were 34, 128, and 949 cpm/mg hemoglobin; those of the HbA1c fraction were 15, 58, and 174 cpm/mg hemoglobin. This evidence of modification of hemoglobin by physiological levels of acetaldehyde from ¹⁴C-label incorporation suggests that an assay more sensitive than chromatographic separation of adducts might be clinically useful in detecting alcoholism or monitoring alcohol detoxification programs.

Eleven male Yucatan miniature swine were subjected to unilateral renal artery constriction (RAC) with an intact contralateral kidney. Five pigs remained at laboratory altitude of 1524 m and served as normoxic, hypertensive controls (NH) while the other six (HH) were subjected to a simulated altitude of 4267 m for 9 weeks to determine the effect of hypoxia on the development of renal hypertension. Systemic blood pressure was increased less in HH as compared with NH. Data from three sib-pairs represented in each treatment group suggested a diverse, apparently familial response to RAC. The hypertensive process, irrespective of treatment groups, included a diminution of free circulating serum thyroxin and an increase in serum sodium. Hypoxia moderates renal hypertension in the miniature pig primarily via a decrement in stroke volume and cardiac output.

The interrelationships among plasma renin activity (PRA, ng AI/ml plasma/hr), aldosterone concentration (ng%), and renal Na⁺-K⁴-ATPase activity (μmole PO₄/mg protein/hr) were studied in 9 weanling normotensive spontaneously hypertensive rats (SHR), 9 adult hypertensive SHR, and 9 weanling and 9 adult normotensive Wistar-Kyoto rats (WKY). All groups were placed on a normal (0.4% sodium) diet. PRA and plasma aldosterone, measured in samples drawn from the ether-anesthetized rat, were higher in weanling SHR (15.2 ± 2.0, 37 ± 4.2) than in WKY. PRA measured in samples collected from a separate group of unanesthetized weanling SHR was also greater than in age-matched WKY. In adult SHR, PRA (6.1 ± 0.9) and plasma aldosterone (20.0 ± 2.7) were decreased. During the weanling period Na⁺-K⁺-ATPase activity in SHR was not only greater than in age-matched WKY but was also increased compared to adult normotensive and hypertensive rats (137 ± 9 weanling SHR, 89 ± 7 weanling WKY, 73 ± 11 adult SHR, 84 ± 17 adult WKY). Thus, during the weanling period the renin-angiotensin-aldosterone (R-A-A) system and renal Na⁺-K⁺-ATPase activity are activated in SHR. The elevation of Na⁺-K⁺-ATPase activity may be due to increased aldosterone levels. It was noted, however, that plasma aldosterone was similar in adult WKY and weanling SHR, while Na⁺-K⁺-ATPase activity was higher in SHR. These findings involving R-A-A and renal Na⁺-K⁺-ATPase activity prior to the elevation of blood pressure suggest that the kidneys may play a role in the initiation of hypertension in SHR.

The present study was designed to determine the effect of folate deficiency on pancreatic acinar cell function. In the first series of experiments, three groups of rats were fed ad libitum regular rat feed, folate-deficient diet, or an equivalent amount of folate-sufficient diet. In the second series of experiments, rats were either fed ad libitum or rendered folate deficient by a purified folate-deficient diet; half of the folate-deficient group was replenished with oral folate. Body weight, pancreatic weight, DNA, [methyl-¹⁴C]thymidine incorporation into DNA, RNA, [8-¹⁴C]adenine incorporation into RNA, protein content, synthesis of proteins, amylase content, and basal and bethanechol-stimulated amylase secretion were determined. The parameters were the same in the rats fed a folate-sufficient diet as in those fed a regular rat feed. Feeding a folate-deficient diet resulted in impaired DNA synthesis as evidenced by diminished incorporation of [methyl-¹⁴C]thymidine into DNA. There was no change in secretion of amylase. Similar results were obtained in the second series of experiments. These studies indicate that folate deficiency (rather than antibiotic content of the diet) impaired pancreatic function. Folate deficiency may therefore contribute to pancreatic injury in malnutrition and alcoholism.

Mice were suspended in a model that simulates weightlessness that occurs during prolonged space flight. After 1 and 2 weeks of suspension in an antiorthostatic (head-down tilt) position, the mice were challenged with polyriboinosinic-polyribocytidylic acid to induce interferon-α/β. Interferon production was severely reduced in mice that had been suspended. When mice were allowed to recover in cages for a week following removal from suspension, they recovered their full interferon-production capacity. Mice suspended in an orthostatic (horizontal) position did not have their interferon production capabilities affected, which indicates that stress per se was not a major component in the effects of antiorthostatic suspension on interferon induction.

Human T cells treated with low levels of interferon (IFN) (1–10 units/ml), and washed to remove the IFN, transferred the same level of antiviral activity to recipient WISH cells as an equivalent IFN treatment alone could induce in WISH cells. Further, when T cells pretreated with IFN (1–10 units/ml) were cocultivated with WISH cells in the presence of IFN (1–10 units/ml), a 2.5- to 5-fold greater level of protection developed than could be expected from the additive effect of each. Antibody to leukocyte, fibroblast, or immune IFN blocked the antiviral effect of the respective IFN types but had no effect on the transfer of antiviral activity initiated by leukocyte, fibroblast, or immune IFN. Also, treatment of T cells with actinomycin D blocked the transfer of antiviral activity of IFN-treated T cells. Taken together, the data suggest that the increased antiviral activity is not merely an additive effect of the IFN, but represents a synergistic amplification of protection most likely due to the combination of the separate effects of IFN and IFN-induced transfer. Such interactions would be expected to play a major role in early protection against virus infections in vivo when low levels of interferon are present and lymphocytes are migrating into the area.

The effects of low (200 ppm) and of high (2000 ppm) ascorbic acid, in a nutritionally adequate diet, on blood ethanol levels have been studied in permanently carotid-cannulated, ethanol-infused, unanesthetized guinea pigs. In the acute study, the postinfusion rate of ethanol decline in the blood of animals treated with ascorbic acid was significantly higher when compared with animals treated with fructose, and the rate in the two treated groups was significantly higher than in untreated controls. In the chronic study, animals were infused with sublethal doses of ethanol (30% of the total caloric intake) for 8 weeks. Blood ethanol levels monitored throughout this period showed, at 3 hr postinfusion, a lower concentration in the group on a high ascorbic acid diet. Both experimental groups receiving ethanol lost significantly more body weight in the second week of dieting; but, while the group on high ascorbic acid regained weight steadily thereafter, the group on low ascorbic acid was still 50 g below the controls at the end of the experiment. Liver, kidney, and adrenal ascorbic acid concentrations were lower in the ethanol-treated groups compared to controls. Examination of the liver revealed more fatty metamorphosis or steatosis in the low ascorbic acid group, but there was no evidence of liver fibrosis or cirrhosis. These results demonstrate the feasibility of utilizing the guinea pig for the study of the biochemical and morphological sequelae of alcoholism. They further support the contention that a diet which is nutritionally adequate may no longer be so in the presence of high ethanol intake, and that supplemental vitamin C ingestion may afford protection against ethanol toxicity.

Seventeen of sixty distal extremities of the thoracic aortas of 12-week-old control male turkeys and 37 of 40 distal extremities of the aortas of turkeys fed 0.07% β-aminopropionitrile (BAPN) from 4 to 12 weeks of age contained areas of cartilaginous metaplasia when examined by light microscopy. The cartilaginous areas were generally elongated and located in the subendothelium of control turkeys, but a roundish area of cartilage was occasionally evident in the deep media. The magnitude of chondroplasia was enhanced by feeding BAPN; the extensive lesion usually extended from the subendothelium to deep in the media. Regardless of treatment, chondrocytes were pleomorphic, contained vacuoles, and had cytoplasmic processes. The cells were separated by pools of proteoglycans and connective tissue. The ultrastructure of chondrocytes in the aortas of both treatment groups was typical of this cell type. They had undulations or projections of the cell membranes. The cisternae of endoplasmic reticulum were dilated and contained electron-translucent material which was similar to extracellular proteoglycans. Golgi apparatus, free ribosomes, mitochondria, glycogen granules, filaments, and a centriole also were present in the cytoplasm. The extracellular matrix, which included collagenous and elastic fibers and also delicate fibrils and interconnecting matrix granules, separated adjacent chondrocytes by spaces of varying size.

Two lymphoblastoid tumor cell lines, the Burkitt lymphoma derived BJAB cell line which is free of Epstein–Barr virus (EBV) and B95–8 cells, which are marmoset lymphocytes transformed by EBV isolated from an infectious mononucleosis patient, were studied in regards to their effects on the blastogenic responsiveness of normal human peripheral blood leukocytes stimulated in vitro with mitogens. Mitomycin C treated tumor cell suspensions, when cocultured with normal human blood leukocytes, markedly depressed the expected blastogenic responses in vitro to concanavalin A, pokeweed mitogen, and phytohemagglutinin. In addition, cell-free sonicates from the cell lines also depressed blastogenic responsiveness of the leukocytes in vitro. Heating the sonicates for 10 min at 100°C markedly diminished the suppressive properties of the sonicates, as did ultraviolet light irradiation. The suppressive activity of the B95–8 sonicates was pelleted by high speed centrifugation as compared to the activity of sonicates derived from the BJAB cells. Further studies are warranted to determine the nature and mechanism of suppression of blastogenic responsiveness of normal human leukocytes by soluble components derived from such lymphoblastoid cell lines.

Cooling of isolated guinea pig tracheal smooth muscle from 38 to 28°C over 2.25 min produced a transient contraction followed by sustained relaxation. The cooling-induced contraction was blocked either by pretreatment with ouabain at concentrations of 10^-5 M or greater or by substitution of normal physiological salt solution with K-free solution. In contrast, the contractile response to cooling was not inhibited by pretreatment with phentolamine (10^-5 M), atropine (10^-5 M), tetrodotoxin (3 × 10^-7 M), diphenhydramine (10^-5 M), cromolyn sodium (10^-3 M), indomethacin (3 × 10^-7 M), nifedipine (10^-7 M), or verapamil (3 × 10^-6 M). Addition of NaHCO₃ to the bath during cooling, preventing a change in pH of the physiological salt solution, did not affect the cooling-induced contraction. It is concluded that cooling of isolated guinea pig trachea produces a transient ouabain-sensitive contraction, and that the data suggest the contraction is mediated by inhibition of Na–K-ATPase in the smooth muscle rather than through neuronal stimulation or chemical mediator release.

Earlier studies from this laboratory suggested that embryonic chick bones in organ culture released into the culture medium a specific inhibitor of bone cell proliferation as defined by inhibition of [³H]TdR incorporation into DNA. Dialysis and membrane ultrafiltration experiments suggested that the inhibitory substance (IS) had a molecular weight between 6000 and 14,000. However, subsequent studies on the purification of IS have revealed that the inhibitory activity in bone-conditioned medium is of lower molecular weight and has several properties in common with thymidine (TdR): (1) IS coeluted with [³H]TdR upon gel filtration chromatography on Sephadex G-10. (2) IS bound to charcoal but not to cation or anion exchange resins. (3) Bone-conditioned medium decreased incorporation of [³H]TdR into the free [³H]TdR pool of cells in monolayer culture. (4) Conditioned medium inhibited [³H]TdR incorporation into [³H]thymidine monophosphate in a reaction catalyzed by thymidine kinase. The equivalent concentration of TdR in conditioned medium as estimated by thymidine kinase assay was sufficient to account for the reduction in [³H]TdR incorporation into bone cell DNA. No evidence was found for a specific inhibitor of bone cell proliferation other than TdR. Hence we conclude that the inhibitory effect of IS is due to dilution of [³H]TdR by nonradioactive TdR. Furthermore, media conditioned by several tumor cell lines also contained a low-molecular-weight component which inhibited [³H]TdR incorporation. The results suggest that organ- and cell-conditioned media can contain significant concentrations of TdR which can artifactually inhibit [³H]TdR incorporation in cell proliferation assays.

In a series of four experiments, the temporal development of acute inhibitory and delayed stimulatory effects of 17β-estradiol (E) on luteinizing hormone (LH) release by superfused rat anterior pituitary cells pulsed with gonadotropin-releasing hormone (GnRH) was studied. Dispersed anterior pituitary cells from ovariectomized rats were cultured on Bio-Beads for 3 days and then placed in columns and superfused for up to 24 hr. During superfusion, the cells were exposed to GnRH pulses (3 × 10^-9 M, one 6-min pulse/hr). Cells treated with E (3 × 10^-10 M) either before (only 24 hr prior to superfusion) or before and during superfusion released significantly (P < 0.05) more LH in response to the first few pulses of GnRH than cells treated with diluent. In contrast, cells treated with E only during superfusion initially released less GnRH-induced LH than cells treated with diluent. In a subsequent experiment, the inhibitory effect of E reached a maximum by 1.5 hr (P < 0.01), and then gradually disappeared after 4.5 hr. Cells superfused simultaneously with E and fixed “low”-dose GnRH (5 × 10^-10 M) pulses did not exhibit enhanced LH responses with time to that dose of GnRH. However, E-superfused cells responded more than diluent-superfused cells to subsequent stimulation with a higher-dose GnRH pulse. Superfusion of cells with E for 16.5 hr in the absence of GnRH pulses also did not increase release of LH to low-dose (5 × 10^-10 M) pulses of GnRH, yet did cause a transitory increase to subsequent high-dose (10^-8 M) GnRH pulses. In conclusion, these results demonstrate the direct biphasic inhibitory then stimulatory effects of E on GnRH-induced LH release by superfused rat anterior pituitary cells. Expression of the stimulatory effect of E is related to the dose of GnRH.

In two experiments, neonatal female BALB/cCrgl or BALB/cfC3HCrgl mice were given subcutaneous injections of 5 μg 17β-estradiol or sesame oil for the first 3 days of life and were ovariectomized at 60 days of age, at which time vaginal concretions (Experiments I and II) or silica (Experiment II) were implanted intravaginally. Mice were examined at 12 months of age. Three abnormal cervicovaginal epithelial responses were noted: persistent vaginal stratification/cornification (PVS); prominent vaginal squamocolumnar junction (SCJ); epithelial pegs, downgrowths, or lesions (dysplasias). PVS, not present in unimplanted controls, occurs in at least half of the members of the neonatally estrogen-treated groups; implants of concretions or silica did not increase its incidence significantly. Although SCJ was observed in implanted but not in unimplanted controls, its incidence was significantly higher in neonatally estrogen-treated mice than in either control group. The elevated incidence in neonatally estrogen-treated mice was not increased further by implantation of concretions or silica. In neonatally estrogenized mice, the subsequent implantation of a concretion significantly increased the incidence of cervicovaginal abnormalities. Increased PVS and SCJ are teratological consequences of neonatal exposure to a small amount of estrogen; on the other hand, increased dysplasias may, in part, be responses of the estrogenized vaginal epithelium to the concretions.

On chronic exposure to hypoxia, the commercially available Hilltop (H) strain of male Sprague–Dawley rats develops severe pulmonary hypertension, right ventricular hypertrophy (RVH), and polycythemia. These signs of chronic mountain sickness are associated with a high mortality rate. In contrast, the Madison (M) strain of Sprague–Dawley rats remains healthy with significantly less severe cardiopulmonary and hematological responses. Breeding experiments under locally controlled conditions were undertaken to determine if the differences between the two strains were genetically determined and to look for possible sex differences. Following 30 to 50 days exposure to a simulated altitude of 18,000 ft, the first generation of male H rats exhibited a higher right ventricular peak systolic pressure (RVPP), a more pronounced RVH, and a greater degree of polycythemia than the male M rats. The H rats had a mortality rate of 40% in contrast to a rate of 0% in the male M rats. The first generation of female H rats also developed a higher RVPP, a greater RVH, and more severe polycythemia than that in the female M rats. There were no differences in RVPP or RVH between the males and females of either strain. Females of both strains tolerated the hypoxic exposure with a 0% mortality rate. The data suggest that the differences between the males of H and M strains in their cardiopulmonary and hematological responses and in their susceptibilities to chronic hypoxia are genetic in nature. They further suggest that the female resistance to hypoxia is not due to milder cardiopulmonary responses. Perhaps female rats tolerate RVH better than male rats, at least of the H strain.

Hollow fiber units containing allogeneic pituitary cells were implanted intracranially into heterozygous (lit/+) and homozygous, mutant (lit/lit) C57BL/6J “little” weanling mice. Over the 48 days of the experiment, heterozygous mice with pituitary cell implants had a lower percentage weight gain than control mice. Homozygous, mutant mice with cell implants, however, made significant weight gains over mutant controls. Long bone lengths were lower, and organ and carcass weights were higher, in heterozygous mice receiving pituitary cell implants than in control mice, but corresponding measurements in mutant mice with and without implants were not significantly different. Supplementation of the diet with thyroid powder increased the percentage weight gain during the latter half of the 48-day period in both genotypes with and without implanted cells. Thyroid-supplemented mutant mice with pituitary cell implants had significantly higher organ and carcass weights than other mutant groups. The little mouse may serve as a model for pituitary studies and for the treatment of isolated growth hormone deficiency type 1 in man.

α-Melanocyte-stimulating hormone (α-MSH) was measured in the mediobasal hypothalamus (MH), median eminence (ME), preoptic-suprachiasmatic area (POA–SCN), anterior (AL), and posterior lobes (PL) of the pituitary gland during the gestational and postpartum periods in the rat. The content of α-MSH in the MH and POA–SCN compared to estrous levels was lower during the later days of gestation and decreased further in the MH during lactation in association with the elevated plasma prolactin (Prl). Distinct increases in the ME content of α-MSH compared to estrous levels occurred on Days 8 and 12 of the gestational period and Day 14 of the postpartum period. A significant increase in PL content of α-MSH compared to Days 5–11 and 17–20 occurred on Day 4 of gestation, and no significant changes were detected in the AP concentration of α-MSH throughout the period studied. In vitro, PLs and ALs from females on Day 4 of gestation secreted more α-MSH into the incubation medium than tissues from animals on Day 20. These results suggest that α-MSH of both brain and pituitary origin may play a role in mediating some of the physiological changes which occur during pregnancy and lactation.

Following termination of vagal stimulation, heart rate increases above control (postvagal tachycardia). This phenomenon has been attributed to vagally mediated release of norepinephrine in the sinus node region, although other contributory factors may be important. The possibility that, during the postvagal period, the chronotropic efficacy of norepinephrine is enhanced was investigated. Mongrel dogs (N = 6) were pretreated with reserpine in order to minimize postvagal tachycardia and hence allow reliable detection of enhanced responsiveness to norepinephrine. The dogs were then anesthetized with chloralose, autonomically decentralized, and instrumented to record electrocardiogram, aortic blood pressure, and electrograms from right atrium and right ventricle. Thirty-, forty-, or sixty-second infusions of norepinephrine were administered via the sinus node artery. The mean cycle length decrease produced by norepinephrine alone was 95 msec (which corresponds to a heart rate increase of +19.6 bpm). After a 30-sec period of vagal stimulation, norepinephrine infusions produced a cycle length decrease of 139 msec (+32.5 bpm). These results are significant at the P < 0.05 level. It is concluded that norepinephrine infusions produce a significantly greater magnitude of tachycardia when administered postvagally. It is proposed that this postvagal potentiation of the chronotropic effect of norepinephrine may contribute to postvagal tachycardia. Indeed, there may be a synergistic relationship between this phenomenon and vagally mediated release of norepinephrine in the mediation of postvagal tachycardia.

Guinea pigs inoculated intravaginally with herpes simplex virus type 2 (HSV-2) strain 1868 produced a serum complement-requiring neutralizing (CRN) antibody during primary acute infection, i.e., 10 days postinoculation. The CRN antibody titers in the guinea pig sera decreased to less than 1:10 after heating at 56°C for 30 min. It was found that 32 units of complement were necessary to obtain a satisfactory HSV-2 neutralizing antibody titer. Nonheated sera significantly reduced virus infectivity titers when mixed with 3.5 log₁₀ PFU of HSV-2 and incubated at 37°C for 20 to 60 min (P < 0.001), whereas the same sera after heating at 56°C for 30 min showed no inhibitory effect. Only 27.3% of infected guinea pigs had low serum non-CRN antibody titers ranging from 1:20 to 1:40. In addition, no evidence of increase in CRN antibody titers was noted during spontaneous recurrent genital herpes infection.

Using specific anti-rat renal renin antibody, the presence of renin in the rat testis was demonstrated by biochemical determination of renin activity. There was no correlation between testicular and plasma renin activity, indicating independent control of testicular and plasma renin levels. Since specific immunohistochemical staining for renin had been observed exclusively in Leydig cells, the effects of hypophysectomy and gonadotropin treatment on the testicular renin were investigated. After hypophysectomy, renin level in the testis decreased significantly, whereas plasma renin was slightly increased. In contrast, testicular renin had remarkably increased through gonadotropin treatment. The results indicate the presence of gonadotropin-dependent renin in the Leydig cells, and suggest a role for it in regulating testicular functions.

The mutagenic character of richterite asbestos was detected in Escherichia coli strain CSH50, using a modified Ames test. Two sets of experimental plates which contained naturally occurring asbestos were used, to one set of which S-9 rat liver homogenate was added. Control plates contained no asbestos. Mutant colonies appeared in significantly greater frequency in both experimental sets as compared with controls, and experimental plates with rat liver homogenate contained the greatest number of mutant colonies. It is postulated that enzymes contained in rat liver metabolize some unknown mutagen introduced with the natural asbestos sample other than the inert richterite fibers themselves. This finding, together with the unlikelihood that bacterial cell walls could be easily penetrated by asbestos fibers, tends to negate fiber penetration as the mechanism of mutagenesis. The mutagenic mechanism operative in E. coli (CSH50) may be applicable to animal cell systems.

Organ distribution and blood concentration profiles were compared following injection of mice with radiolabeled test agents via the lateral tail vein or retroorbital venous sinus. Monoclonal antibodies directed against B16 melanoma of C57BL/6 origin were labeled with iodine-125. Thymocytes from BALB/c mice and B16 melanoma cells were labeled with technetium-99m sodium pertechnetate (Na ^99mTcO₄). Animals were injected with 5 μCi of iodinated antibody, 5 × 10⁵ syngeneic thymocytes, 2.5 × 10⁵ melanoma cells, or 10 μCi Na ^99mTcO₄ in 0.2 ml saline via either route. In non-tumor-bearing C57BL/6 mice radiolabeled monoclonal antibody was found primarily in the gastrointestinal tract, liver, and blood. Na ^99mTcO₄ localized in the gastrointestinal tract, ^99mTc-labeled thymocytes in the spleen and liver, and ^99mTc-labeled B16 melanoma cells in the liver and lungs. Pharmacokinetic analysis of blood samples taken 4, 8, and 12 min following injection of the labeled agents suggested that the iodinated antibody had less vascular permeability than Na ^99mTcO₄ and that thymocytes and B16 melanoma cells were trapped in the pulmonary vasculature as they passed through the lungs. It is noteworthy that no biologically significant differences in organ distribution patterns or blood decay profiles were found between lateral tail vein and retroorbital routes. The data clearly indicate that these routes can be used interchangeably with one another for intravenous injections.

The purpose of the present study was to examine the effect of administering an opiate receptor antagonist, naltrexone (NALT) on the decline in pituitary thyrotropin (TSH) release induced by both acute and chronic stress, and to determine whether norepinephrine (NE) is involved in the mechanism by which opiate receptor blockade counteracts inhibition of TSH release during stress. Administration of NALT, a specific opiate receptor antagonist, significantly attenuated the decrease in plasma TSH observed after acute exposure to ether or restraint stress. The ability of NALT to prevent suppression of circulating TSH in ether-stressed rats was blocked by pharmacological suppression of NE activity induced by pretreatment with diethyldithiocarbamate (DDC) or phenoxybenzamine (PB), both NE antagonists. In chronically stressed rats, thrice daily injections of NALT attenuated the sustained decline in circulating TSH, and resulted in a significant elevation in plasma TSH when compared with stressed, saline-treated animals. Pretreatment with DDC prior to NALT injection abolished this stimulatory effect of NALT. These observations indicate that opiate/receptor interaction is prerequisite for the decrease in circulating TSH release during both acute and chronic stress, and support the hypothesis that endogenous opioid peptides (EOPs) mediate the suppressive effect of stress on TSH release. The finding that uninterrupted NE function is necessary for NALT's action on TSH release during stress suggests that the suppressive effect of stress on TSH and its reversal by opiate antagonists involves alterations in hypothalamic NE activity.

Milk was collected for the first 21 days of lactation twice daily from dairy cows and once daily from goats, sheep, and guinea pigs. Thyroxine (T₄) and triiodothyronine (T₃) were extracted from 100 μl of milk using acidified ethanol. T₄ and T₃ were reconstituted in 100 μl buffer and measured by radioimmunoassay. Concentrations (ng/ml) of T₄ and T₃ for milk of cows, goats, sheep, and guinea pigs, respectively, were: 0.97 and 0.94, 1.24 and 0.52, 0.99 and 0.79, and 1.41 and 0.53. T₄ concentration for guinea pig milk was significantly higher than for cow and sheep milk, but not for goat milk (P < 0.05). T₃ was found in higher concentration in milk of cows and sheep than in milk of goats and guinea pigs (P < 0.05). Species differences in conversion of T₄ to T₃ in mammary gland cells are suggested. Summations of T₄ and T₃ concentrations in milk indicated no differences among the four species. Regression analyses of changes in milk production, T₄ and T₃ concentrations, total T₄ and T₃ in milk per day, and ratios of T₄ to T₃ revealed variations in patterns. Concentrations of T₄ or T₃ tended to decrease as lactation progressed over 21 days. Total T₃ tended to increase, and the ratio of T₄ to T₃ tended to decrease. Amounts of T₄ and T₃ available to offspring from milk were calculated to be minor sources (4 to 7%) of total requirements for maintenance of metabolic functions.

Short-chain fatty acids (SCFA) produced during fermentation of plant fibers and absorbed from the colon may affect hepatic glucose metabolism. We examined the effects of different fatty acids on rates of glucose production and glycolysis in isolated rat hepatocytes. Acetate, butyrate, and long-chain fatty acids significantly increased glucose production from lactate. However, propionate and valerate significantly decreased glucose production from lactate. Whereas 5 mM butyrate increased the incorporation of [¹⁴C]lactate into [¹⁴C]glucose by 80%, 5 mM propionate produced a 67% decrease. Glycolysis was significantly decreased by acetate, butyrate, and long-chain fatty acids. However, propionate and valerate significantly increased glycolysis. Thus propionate, which inhibits hepatic acetate metabolism, acts to increase glucose use and decrease glucose production. Plant fibers may influence hepatic glucose metabolism via their SCFA metabolites.

Angiotensin II has been previously implicated as a mediator of vasoconstriction during the development of hypoxic pulmonary hypertension. The effect of angiotensin-converting enzyme inhibition with teprotide (SQ 20881) on development of pulmonary hypertension was determined by measurement of the drug's ability to modify hypertension-induced protein synthetic changes in the rat pulmonary trunk. Rats were injected with either SQ 20881 (2 mg/kg body wt every 8 hr) or saline vehicle during exposure to chronic hypoxia at 0.5 atm for either 3 or 7 days. Comparisons were made of tissue weight, absolute protein content, and in vitro synthesis of collagen and noncollagen protein of the pulmonary trunks of SQ-treated hypoxic, SQ-treated normoxic, saline-treated hypoxic, and saline-treated normoxic rats. Treatment of hypoxic rats with SQ 20881 was found to significantly decrease right ventricular pressure, tissue weight, absolute protein content, and in vitro protein synthesis after 7 days compared to saline-treated hypoxic rats. Neither right ventricular hypertrophy nor the development of polycythemia was decreased by SQ 20881 treatment.