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The influence of sex steroid hormones on ganglioside biosynthesis in rat kidney.

Castrated male rats were subcutaneously injected testosterone (5 and 10 mg) or a mixture of beta-estradiol and progesterone (1 microg + 2 mg), to determine whether sex steroid hormones (testosterone, beta-estradiol, progesterone) might affect the content of sialoglycoproteins, the content and pattern of lipid-bound sialic acid, and the activities of sialyltransferase (SAT) I and SAT II in the Golgi-rich membrane fraction isolated from rat kidney. During four days testosterone did not affect significantly the content of proteins, sialoglycoproteins and total gangliosides, but increased the content of b-series gangliosides from 0.05 +/- 0.006 (untreated animals injected subcutaneously with 0.1 ml DMSO for four days) to 0.16 +/- 0.02 nmol sialic acid (SA) per mg protein (castrated animals injected subcutaneously with 10 mg testosterone/0.1 ml DMSO for four days). This increase was due to the increase in GD3 ganglioside from 0.03 to 0.12 nmol SA/mg protein, and to the decrease of GM3 ganglioside from 0.06 to 0.03 nmol SA/mg protein by testosterone administration. The major ganglioside in the rat kidney was GM3, constituting 63% (control group) and 51% (castrated animals injected daily with 10 mg testosterone) of all gangliosides. Castration itself induced an increase in the rat kidney SAT I and SAT II activities from 712 +/- 130 to 1723 +/- 412 pmol/h x mg protein and from 208 +/- 48 to 751 +/- 176 pmol/h x mg protein, respectively. However, subsequent administration of testosterone, at the highest concentration tested, reversed this effect. In the kidneys of castrated rats, a mixture of beta-estradiol and progesterone decreased SAT II activity from 208 +/- 48 to 87 +/- 33 pmol/h x mg protein.

Animals↗

Comparison of the ability of recombinant human parathyroid hormone, rhPTH-(1-84), and hPTH-(1-31)NH2 to stimulate femoral trabecular bone growth in ovariectomized rats.

A recombinant human parathyroid hormone, rhPTH-(1-84), which is currently in Phase II clinical trial, and hPTH-(1-31)NH2 (Ostabolin) are promising anabolic agents for treating osteoporosis because they can stimulate cortical and trabecular bone growth in osteopenic, ovariectomized (OVX) rats and in osteoporotic, postmenopausal women when injected subcutaneously and intermittently at low doses. We have now found that, despite their different sizes and signaling properties (rhPTH-(1-84) stimulates adenylyl cyclase and phospholipase C; hPTH-(1-31)NH2 only stimulates adenylyl cyclase), they are equally osteogenic in OVX rats. Thus daily subcutaneous injections of 0.6 nmol/100 g of body weight of rhPTH-(1-84) or hPTH-(1-31)NH2 into 3-month-old OVX rats for 6 weeks starting 2 weeks after OVX equally reduced the otherwise large OVX-triggered loss of femoral trabecular bone. Daily subcutaneous injections of 0. 4 or 0.8 nmol/100 g of body weight of the two agents for 6 weeks also equally increased the mean thickness of the remaining femoral trabeculae in 3-month-old and 1-year-old OVX rats to 20 to 80% above the value in normal animals when started 9 weeks after ovariectomy.

Animals↗

[Establishment of human hepatocellular carcinoma cell line with spontaneous pulmonary metastasis through in vivo selection].

OBJECTIVE: To establish a hepatocellular carcinoma (HCC) cell line from lung metastatic lesions of human HCC in nude mice so as to provide suitable model for the study of lung-metastasis-related molecular mechanisms. METHODS: HCC clone cells MHCC97-H were inoculated subcutaneously into nude mice. The pulmonary metastatic lesions were harvested from the moribund animals and then re-implanted into the nude mice for the second round of selection. The same procedure was repeated twice. New cell line from the third round of lung metastasis was thus established and the following parameters were studied: morphology, in vitro growth curve, plate efficiency, migration, karyotype, flow cytometry, immunocytochemistry for AFP, albumin and cytokeratin 8 (CK8), flourescent PCR for HBV DNA, tumorigenicity and spontaneous metastasis via subcutaneous injection and orthotopic implantation of tumor tissue. RESULTS: The cell line established from nude mouse lung metastasis was designated as HCCLM3, consisting of polygonal epithelial cells with hypotriploid karyotype, its main range of chromosome being 55 - 58. The cell population doubling time was 34.9 hours, plate efficiency 32.4 +/- 3.2%, and cell migration rate 20 +/- 2 microm/h. The cells were positive for AFP, albumin, CK8, and P16 and negative for P53, nm23 and HBsAg. Fluorescent PCR showed HBV DNA integration into cellular genome. When 5 x 10(6) cells were injected subcutaneously into nude mice, the tumorigenicity was 100% with a latency period of 11 +/- 1 d. Five weeks after subcutaneous injection, the pulmonary metastatic rate was 100%, the median number of lung metastases being 121 per mouse. After orthotopic implantation of tumor tissue into nude mouse liver for 35 d, wide spread regional and distant metastases occurred, the metastatic rate was 100% for abdominal wall, 80% for organs in abdominal cavity, 100% for liver, 70% for diaphragm, and 100% for lung. The median number of lung metastatic lesions was 268 per mouse. CONCLUSION: A new HCC cell line has been established, which is characterized by high pulmonary metastasis via both subcutaneous and orthotopic inoculation. It provides a new model for the study of liver cancer metastasis.

Animals↗

A reproducible model for the induction of avian cellulitis in broiler chickens.

Avian cellulitis was reproduced in 39-day-old broilers by subcutaneous injection of Escherichia coli originally isolated from a cellulitis lesion. One hundred percent of the birds injected with the isolate on the dorsal and ventral surfaces developed characteristic fibrino-caseous plaques. A slightly lower percentage (90%) of the birds injected subcutaneously in the inguinal area developed the same lesions. Only 30% of the birds that had been inoculated by scratching the skin and swabbing the bacterial inoculum onto the wound developed the lesion. No birds inoculated by swabbing the inoculum onto a feather follicle, from which the feather had been pulled, developed cellulitis. Characteristic cellulitis plaques could be produced as early as 18 hr postinfection (PI). Lesions, consisting of a serosanguinous, yellow-pink-to-orange-tinged fluid appeared as early as 6 hr PI. The lesions progressed, changing to a more thin, yellow, purulent fluid by 12 hr PI followed by plaque formation. Although there was a trend for lesion size to diminish with time, the majority of the challenged birds, examined as late as 3 wk PI, still had prominent cellulitis plaques. Lesions in birds injected subcutaneously on the dorsal surface sometimes extended into other regions of the body, including the abdominal region, and thereby resembled the type of lesions that have previously been described as type I or hatchery-borne cellulitis.

Animals↗

Effect of solvents on methylcholanthrene-induced carcinogenesis in mice.

We have shown previously, with X-chromosome linked phosphoglycerate kinase (PGK) cell markers, that the fibrosarcomas induced in mice by subcutaneous injection of 0.2-2.0 mg of methylcholanthrene (MCA) dissolved in olive oil had a multicellular origin. In this study we used a constant dose of MCA injected subcutaneously and compared the effects on tumorigenesis of using benzene or olive oil as the solvents. The benzene and olive oil groups differed in tumor incidence (100% versus 50%), location (muscle versus skin), and type (rhabdomyosarcomas versus fibrosarcomas). Each of the nine olive-oil MCA-induced tumors showed both PGK enzyme types, whereas 13 ot 16 tumors induced by MCA in benzene displayed only a single PGK type. This difference in PGK phenotypes suggests that the MCA-benzene neoplasms arose from one or a few cells. We conclude from these results that the nature of the solvent plays an important role in the type and number of cells from which tumors induced by subcutaneous injection of MCA arise.

Animals↗

Simultaneous observation of endocrine and exocrine functions of the pancreas responding to somatostatin in man.

Six patients who underwent segmental autotransplantation of the caudal pancreas (SAT) following total pancreatectomy for pancreatic cancer were investigated. The graft was transplanted to the left groin, and pancreatic juice was diverted outside through a polyethylene tube indwelled into the main pancreatic duct. In these SAT patients, the responses of insulin (IRIS) in terms of plasma levels and pancreatic secretion to subcutaneous injections of somatostatin octreotide (Sandostatin: SMS201-995) were simultaneously observed. Four doses (0.039, 0.156, 0.625 and 2.5 micrograms/kg) of SMS201-995 were given on separate days. As a control, saline was injected subcutaneously. Standard liquid test meal was given 1 h after the subcutaneous injection. The basal plasma IRI were significantly decreased with doses greater than 0.156 microgram/kg. The postprandial responses of IRI was also significantly suppressed with the same doses. On the other hand, the basal pancreatic exocrine secretion was significantly suppressed with doses greater than 0.625 microgram/kg. The postprandial pancreatic exocrine secretion was also significantly suppressed with doses greater than 0.625 microgram/kg. Those suppressions were dose-dependent. The postprandial CCK secretion was also significantly suppressed in dose-dependent manner with SMS201-995. The CCK suppression was significantly correlated with the suppression of pancreatic exocrine secretion. This clinical study under the setting of SAT demonstrated not only the direct inhibitory effect of somatostatin on both the islet and acinar cells but also, probably, the indirect inhibitory effect on the acinal cells via suppression of CCK release in humans.

Aged↗

Artificial induction of lactation in ewes: the role of prolactin.

The mammary glands of 30 non-pregnant, intact ewes were developed by subcutaneously injecting oestrogen plus progesterone at intervals of 3 days from day 0 to day 27. Two days later (day 29), 15 ewes were injected subcutaneously with 18 mg ergocryptine, to inhibit specifically secretion of prolactin. Then groups of ewes, each comprising five ergocryptiine-treated and five untreated ewes, were injected from days 30 to 34 with either four intravenous injections each day of 1 i.u. syntocinon, one subcutaneous injection each day of 10 mg dexamethasone trimethylacetate, or two subcutaneous injections each day of 2-5 mg oestradiol benzoate plus 6-25 mg progesterone. All ewes were milked by hand on days 30-50. Within 24 h of injecting ergocryptine, levels of prolactin in serum were reduced to negligible values (less than 2 ng/ml). Comparison of results for ewes not receiving ergocryptine showed that syntocinon, dexamethasone and oestradiol benzoate plus progesterone, at the doses used, were equally effective in initiating milk secretion. Peak yields of 0-23-0-27 kg/day were achieved. On the other hand, ewes treated with ergocryptine before syntocinon or dexamethasone produced peak yields of only 0-12-0-13 kg/day and ewes treated with ergocryptine before oestradiol benzoate plus progesterone produced negligible amounts of secretion. The results suggest that syntocinon and dexamethasone were either lactogenic per se or effected the release of hormones of the lactogenic complex other than prolactin. However, oestradiol benzoate plus progesterone appeared to be lactogenic by virtue of the influence of oestrogen on the secretion of prolactin.

Animals↗

Short-term macrophage interleukin-1 response to injected silicone in a rat model.

The short-term effects of silicone particles on the ability of splenic and peritoneal macrophages to produce Interleukin-1 (IL-1) were assessed. Lewis rats were divided into three groups. Group 1 animals (n=12) were injected subcutaneously with 2.5 ml of sterile saline; Group 2 animals (n=12) received 2.5 ml of Freund's Complete Adjuvant (FCA); Group 3 animals (n=12) received subcutaneous injections (2.5 ml) of a sonicated slurry comprised of equal parts of FCA and silicone. At days 1, 4, and 7 single-cell suspensions of splenic (10(8) cells/ml) and peritoneal (10(6) cells/ml) macrophages of four animals from each group were prepared. The supernatants from macrophages cultured in the presence or absence of lipopolysaccharide (LPS) were added to IL-1 dependent mouse thymocytes for 72 hrs, and subsequently pulsed with 3H-thymidine. Radioactivity incorporation was determined after 18 hrs. Peritoneal macrophage culture supernatants had significantly higher activity than splenic macrophages and all macrophages stimulated with LPS had significantly higher activity. At days 1 and 7, there were no significant differences in 3H-thymidine uptake. At day 4 there was an elevation of 3H-thymidine uptake from supernatant of peritoneal macrophages treated with FCA/silicone suggesting subcutaneous silicone injection has short-term effects.

Animals↗

The effect of subcutaneously administered terbutaline on serum potassium in asymptomatic adult asthmatics.

Several recent reports have linked the administration of beta-adrenergic agents to the development of hypokalemia. This led us to study, under controlled conditions, the effects of a subcutaneous injection of 0.25 mg terbutaline sulfate on serum potassium (K+), sodium (Na+), glucose, and insulin in a group of asymptomatic asthmatics. Heart rate (HR), blood pressure (BP), and forced expiratory volume in one second (FEV) were measured as parameters of beta-adrenergic stimulation. Each of 7 subjects was studied on 2 separate mornings after an overnight fast. Measurements were obtained before and at various time intervals after 0.25 mg) of terbutaline sulfate or 0.25 ml of a normal saline solution (placebo) were injected subcutaneously in a double-blind randomized crossover fashion. Terbutaline injection significantly increased HR and FEV and lowered diastolic BP for a period in excess of 3 h, indicating a prolonged beta stimulatory action of the drug. Significant effects on glucose, K+, and insulin were observed as early as 15 min after terbutaline injection; serum glucose and insulin increased and remained elevated for at least 2 h. Serum K+ fell significantly, reached a peak decline of 14% from the control value at 30 min after the injection and remained decreased for a period in excess of after subcutaneous terbutaline injection should be interpreted in the light of these effects of the drug and that terbutaline should be used with caution in patients prone to hypokalemia.

Adult↗

Comparison of transthecal and subcutaneous single-injection digital block techniques.

A randomized double-blinded study was performed on 20 normal volunteers to evaluate 2 different techniques of single-injection digital anesthesia. Single-injection transthecal digital block technique was used to anesthetize 1 index finger and single-injection subcutaneous technique to block the other index finger. Pain and light touch were evaluated and sensory nerve-conduction studies were performed on both index fingers. These data were obtained prior to the nerve blocks and then at 10-minute intervals until recovery from the anesthesia. The method of anesthesia was found to have no effect on the distribution, onset, and duration of anesthesia. Median and radial nerve sensory nerve action potential amplitude reductions following digital anesthesia were also not influenced by the technique of anesthesia. Single-injection subcutaneous block was found to be easier to administer and to produce less pain during and 24 hours after injection than did the single-injection transthecal technique.

Adult↗

[Abortive activity of mimosine and its prevention (author's transl)].

18 out of 19 female mice treated orally with different levels of mimosine in the drinking water were fertile. Subcutaneous injection of mimosine (400 mg/kg) to 23 pregnant mice, in a single dose on days 5, 10, 15 or 17, resulted in death and resorption or expulsion of dead embryos, except in one, early inoculated. L-Phenylalanine (600 mg/kg) injected subcutaneously on day 17 prevented abortion in 4 out of 5 mice subsequently injected with mimosine.

Abortifacient Agents↗

Toxicity during early development of the mouse nervous system of a scorpion neurotoxin active on sodium channels.

The lethal effects of scorpion envenomation is due to neurotoxins active on voltage-sensitive sodium channels. Dysfunctions of the peripheral and central nervous systems with neurological manifestations are commonly observed after scorpion stings, specially in young children. Since the neurotoxicity of venom fraction is greatly higher by intracerebroventricular than by subcutaneous injections, a direct effect of venom on CNS cannot be excluded specially in infants where the blood-brain barrier is not fully functional. We investigated the activity of a neurotoxin from the scorpion Androctonus australis hector (AahII) in newborn mice at 3, 7 and 14 days after birth and in adults. Young mice (P3, P7) were more sensitive to AahII injected subcutaneously than were adults, but were less sensitive to intracerebroventricular injection. The affinity of AahII for its receptor site on brain synaptosomes from P3 and P7 mice was slightly higher and the density of the binding sites was half that of adult mice. After subcutaneous injection of [125I]-AahII it was also observed that a small amount of radioactivity was found in brains of neonate mice but not in that of adults. This amount is however extremely lower than the value of the LD50 determined by intracerebroventricular injection. Results are consistent with a peripheral action of AahII and show that its toxic activity changes during the mouse nervous system development.

Age Factors↗

[Study of the methods of specific desensitization therapy in immediate hypersensitivity to Streptococcus under experimental conditions. 1. Immunological data].

Three series of experiments were conducted on 427 guinea pigs. A model of allergy of the immediate type was obtained by 3-fold subcutaneous injections of 2 mg of lysed streptoallergen with an imcomplete Freund's adjuvant. The effect of allergens (corpuscular--vaccines, lysed, and streptoallergens after Ando-Verzhikovsky) varying by physico-chemical properties was studied in the first experimental series. The best hyposensitizing action was produced by vaccine used for the study of the influence of various doses on the sensitized organism. Two doses were approved: the threshold one (diluted 10 times) and the subthreshold one (diluted 10000 times). The use of the threshold doses caused reduction of increased sensitivity of the immediate type. In the III experimental series this dose was injected subcutaneously, intradermally, and intravenously. Subcutaneous method proved to produce a more marked hyposensitizing action in comparison with other methods.

Allergens↗

Biotolerance of a semisolid hydrophobic biodegradable poly(ortho ester) for controlled drug delivery.

We evaluated the biotolerance of a new semisolid poly(ortho ester) (POE) intended for controlled drug delivery. Two different investigations were carried out in rats: subcutaneous injections and the cage-implant system. Sodium hyaluronate (SH), 1%, in phosphate buffer was used as a noninflammatory control. When injected subcutaneously in rats, the POE induced a mild and local inflammation at 3 and 7 days followed by a minimal chronic inflammation at 14 and 21 days. When using the cage-implant system, we quantified the inflammatory components of the exudate surrounding the implanted material within the cage system at 3, 7, 14, and 21 days. The total leukocyte and macrophage concentrations were higher only at 7 days for both SH and POE compared to the control (empty cage). The other parameters were not significantly different from the control. These results show that POE is well-tolerated by rats when used subcutaneously.

Animals↗

Monoclonal antibodies in the lymphatics: selective delivery to lymph node metastases of a solid tumor.

After subcutaneous injection, monoclonal antibodies directed against a tumor can enter local lymphatic vessels, pass to the draining lymph nodes, and bind to metastases there. Lymphatic delivery of antibody to early metastases is more efficient than intravenous administration, and the lymphatic route can be used to image smaller metastatic deposits. Perhaps more important, the lymphatic route minimizes binding of antibodies to circulating tumor antigens and to cross-reactive antigens present on normal tissues. Antibodies inappropriate for intravenous use because of binding to normal tissues may therefore be useful against lymph node metastases when injected subcutaneously or directly into lymphatic vessels.

Animals↗

Effects of subcutaneous calcitriol administration on plasma calcium and parathyroid hormone concentrations in continuous ambulatory peritoneal dialysis uremic patients.

OBJECTIVE: To ascertain whether the parathyroid hormone (PTH) secretion of continuous ambulatory peritoneal dialysis (CAPD) uremic patients could be suppressed by repeated subcutaneous injections of calcitriol (CLT). DESIGN: Nonrandomized prospective study with weekly evaluation. SETTING: Hospital CAPD clinic. PATIENTS: Seven uremic CAPD patients with signs of severe hyperparathyroidism. INTERVENTIONS: Patients were treated with CLT (2 micrograms), injected subcutaneously three times a week, on alternate days over a period of 8 weeks. MEASUREMENTS: Plasma PTH, ionized calcium (Ca), serum phosphate (Pi), and alkaline phosphatase (AP) were assayed before the start of CLT therapy and weekly thereafter. RESULTS: The average basal PTH was 349 +/- 26 pg/mL (mean +/- SD). It fell significantly by the fifth week to 158 +/- 20, then leveled off. Analysis of the individual data, however, revealed that only 5 of 7 patients had a significant decrease in plasma PTH. Basal Ca was +/- .02 mmol/L; it increased continuously throughout the study, significantly by the fourth week, reaching a level of 1.33 +/- 0.3 mmol/L at the sixth week, then declined slightly. In those patients with significantly decreased PTH, there was an inverse correlation between PTH and the corresponding Ca levels. CONCLUSIONS: In some CAPD patients subcutaneous administration of CLT significantly suppresses PTH. This effect is mainly mediated via an increase in ionized calcium, but a direct inhibitory effect of the vitamin on parathyroid glands cannot be excluded.

Adult↗

Erythema multiforme associated human autoantibodies against desmoplakin I and II: biochemical characterization and passive transfer studies into newborn mice.

The demonstration of circulating autoantibodies directed against the constitutive desmosomal plaque proteins desmoplakin (dp) I and II in mucocutaneous lesions in a subset of patients with erythema multiforme major, suggests that humoral immune mechanisms may play a role in the pathogenesis of this severe skin disease. In this study we identified a specific peptide sequence--YSYSYS--representing an antigenic binding site for the human autoantibodies. This epitope is localized at the extreme carboxy terminal domain of dp thought to be responsible for the assembly of keratin filaments with desmosomes. To test the possibility whether these antibodies may exert any pathologic effects in vivo, human autoantibodies were affinity purified on a corresponding synthetic peptide matrix and peptide-specific antibodies were raised in rabbits. After repeated subcutaneous injections into newborn mice, affinity-purified human autoantibodies and anti-peptide rabbit IgG were detected on desmosomal plaques of keratinocytes overlying the injection site. Histologic and electron microscopic examinations showed hydropic degeneration of basal and suprabasal keratinocytes, dyskeratosis, signs of suprabasal acantholysis, and keratin filaments detached from the desmosomal plaques clumping around the nucleus. We demonstrate that autoantibodies are directed to an epitope within a dp domain crucial for the interaction of keratin filaments with desmosomes, and, when injected subcutaneously into newborn mice, produce pathologic changes. These findings imply that autoantibodies to dp could impair the function of desmosome-keratin filament complexes suggesting a pathogenic role in vivo.

Animals↗