Intraperitoneal Injection Routes Research Articles

Efficacy of Chitosan-PLGA encapsulated trivalent oral vaccine against viral haemorrhagic septicemia virus, Streptococcus parauberis, and Miamiensis avidus in olive flounder (Paralichthys olivaceus)

The present study was conducted to assess the protective efficacy of a trivalent oral vaccine containing chitosan–PLGA encapsulated inactivated viral haemorrhagic septicemia virus (VHSV), Streptococcus parauberis serotype I and Miamiensis avidus antigens, followed by its oral (incorporated in feed) administration to olive flounder (Paralichthys olivaceus) fingerlings for a period of 15-consecutive days. After 35 days of initial vaccination, three separate challenge studies were conducted at the optimal temperature of the targeted pathogens using an intraperitoneal injection route. RPS analysis revealed moderate protection in the immunized group against all the three pathogens viz., VHSV (53.30% RPS), S. parauberis serotype-I (33.30% RPS), and M. avidus (66.75% RPS), as compared to the respective non-vaccinated challenge (NVC) control group. In addition, the immunized fish demonstrated significantly (p < 0.05) higher specific antibody titres in serum and significant (p < 0.05) upregulation in the transcript levels of immune genes of Igs (IgM, IgT, pIgR), TLRs (TLR 2, TLR 7), cytokines (IL-1β, IL-8) and complement pathway (C3) in the mucosal and systemic tissues than those of NVC control fish, suggesting orchestration of pathogen-specific host immune responses thereby favouring its combativeness against the three pathogens. The expression dynamics of IFN-γ, Mx, caspase 3 genes post VHSV challenge; IFN-γ, TLR 2, caspase 1 genes post S. parauberis serotype I challenge and CD-8α, IL-10, TNF-α genes post M. avidus challenge further substantiates the efficacy of the vaccine in stimulating antiviral, antibacterial and antiparasitic immune responses in the host resulting in their better survival. The findings from the present study reflect that the formulated trivalent oral vaccine incorporating VHSV, S. parauberis serotype I and M. avidus antigens can be a promising prophylactic strategy to prevent the associated disease outbreaks in olive flounder.

Effects of administration route on uptake kinetics of 18F-sodium fluoride positron emission tomography in mice

18F-sodium fluoride (18F-NaF) is a positron emission tomography (PET) radiotracer widely used in skeletal imaging and has also been proposed as a biomarker of active calcification in atherosclerosis. Like most PET radiotracers, 18F-NaF is typically administered intravenously. However in small animal research intravenous administrations can be challenging, because partial paravenous injection is common due to the small calibre of the superficial tail veins and repeat administrations via tail veins can lead to tissue injury therefore limiting the total number of longitudinal scanning points. In this paper, the feasibility of using intra-peritoneal route of injection of 8F-NaF to study calcification in mice was studied by looking at the kinetic and uptake profiles of normal soft tissues and bones versus intra-vascular injections. Dynamic PET was performed for 60 min on nineteen isoflurane-anesthetized male Swiss mice after femoral artery (n = 7), femoral vein (n = 6) or intraperitoneal (n = 6) injection of 8F-NaF. PET data were reconstructed and the standardised uptake value (SUV) and standardised uptake value ratio (SUVr) were estimated from the last three frames between 45- and 60-min and 8F-NaF uptake constant (Ki) was derived by Patlak graphical analysis. In soft tissue, the 18F-NaF perfusion phase changes depending on the type on injection route, whereas the uptake phase is similar regardless of the administration route. In bone tissue SUV, SUVr and Ki measures were not significantly different between the three administration routes. Comparison between PET and CT measures showed that bones that had the highest CT density displayed the lowest PET activity and conversely, bones where CT units were low had high 8F-NaF uptake. Intraperitoneal injection is a valid and practical alternative to the intra-vascular injections in small-animal 18F-NaF PET imaging providing equivalent pharmacokinetic data. CT outcome measures report on sites of stablished calcification whereas PET measures sites of higher complexity and active calcification.

Prophylactic antibiotic usage by Belgian veterinarians during elective caesarean section in Belgian blue cattle

The use of prophylactic antibiotics in veterinary surgery reduces the occurrence of postoperative complications. In order to limit the development of antimicrobial resistance in bacteria and to maximize therapy efficacy, antibiotics should be used prudently and efficiently. A survey was sent to Wallonian rural veterinarians in order to assess the use of antibiotics before, during and after bovine elective caesarean section, the most common surgical procedure in Belgian rural practice. Results were analysed in the light of the current guidelines formulated by the Belgian Centre of Expertise on Antimicrobial Consumption and Resistance in Animals.Among 380 contacted veterinarians, 113 answered the survey. All veterinarians use antibiotics during caesarean section. Veterinarians predominantly use penicillin as the first drug of choice, whereas a minority use drugs considered as second or third choice, such as amoxicillin, oxytetracyclin or lincomycin-spectinomycin. Also, 31/113 veterinarians simultaneously use molecules from different antibiotic classes. The majority (100/113) of veterinarians inject antibiotics during or after surgery, while a minority (13/113) administer antibiotics preoperatively. Most veterinarians (67/113) limit the duration of their antibiotic treatment to 1 day. Considering the administration route, most veterinarians (88/113) systematically use the intraperitoneal injection route, which is not registered. Intramuscular injection and injection between muscle layers during surgery are carried out by 82/113 and 43/113 respondents, respectively. Twenty-two respondents apply intra-uterine antibiotics. Most veterinarians (87/113) combine several administration routes. The dosage of antibiotics varies enormously and excessive injection volumes are common, especially when multiple injection routes are combined.Our results show a striking lack of consistency in the antibiotic therapy during elective CS by rural veterinarians. Whereas the drug of choice and the treatment duration were largely in line with the current guidelines, this is certainly not the case for the dosage and the administration route. The intraperitoneal injection of antibiotics cannot be justified. Incorrect dosage of antibiotics should be avoided at all cost. The use of second and third line molecules and the simultaneous use of different antibiotic classes should be discouraged. A major challenge lies in the education of veterinary students and the sensitization of practitioners to avoid or unlearn unnecessary habits concerning antibiotic use. Finally, more clinical trials are needed in order to refine the current guidelines for antibiotic use and to determine the optimal drug of choice, treatment moment and application route in elective caesarean section in cattle.

Histopathological Study on The Effect of Neuregulin-1 Against Cardiotoxicity Induced By Trastuzumab in Adult Mice

Objective: This work carried out to investigate the potential therapeutic effect of Neuregulin-1 against cardiotoxicity induced by trastuzumab in male mice. Materials and Methods: Forty five adult male and female mice (weighing 35-50 gm) were divided into 3 groups randomly, each group contains 15 animals, group A which received distilled water as control by using intraperitoneal injection (IP) route for 10 days group B, received Trastuzumab as 6 mg/kg/day by using intraperitoneal injection route for 10 days while group C, received Neuregulin-1 (2.5 mg/day/animal) just before the administration of Trastuzumab (6mg/kg/day) given by IP route for 10 days. The significance of difference were tested by using ANOVA followed by Tukey test. Statistical significance depends on the P&lt;0.05. Results: The histological score of cardiac tissue changes showed that a highly increase in mice administrated with trastuzumab when compared to control group while pretreatment with Neuregulin-1 , made a decrease in histological changing score when compared with trastuzumab group. The percentage of immunohistochemical score of stain uptake for TNF-α and IL-1β expression in cardiac tissue shows a highly increase in mice administrated when compared with control group, furthermore, pretreated with neuregulin-1 showed a decrease in the percentage of staining when compared with trastuzumab group. Conclusion: Neuregulin-1 can be used as a cardioprotective in cardiac complications of cancer treatment by its ability to attenuate the cardiotoxic effect of trastuzumab by decreasing in histological changing score and decrease in the percentage of staining when compared with trastuzumab group.

Histopathological Study on The Effect of Neuregulin-1 Against Cardiotoxicity Induced By Trastuzumab in Adult Mice

Objective: This work carried out to investigate the potential therapeutic effect of&#x0D; Neuregulin-1 against cardiotoxicity induced by trastuzumab in male mice.&#x0D; Materials and Methods: Forty five adult male and female mice (weighing 35-50 gm) were divided into 3 groups randomly, each group contains 15 animals, group A which received distilled water as control by using intraperitoneal injection (IP) route for 10 days group B, received Trastuzumab as 6 mg/kg/day by using intraperitoneal injection route for 10 days while group C, received Neuregulin-1 (2.5 mg/day/animal) just before the administration of Trastuzumab (6mg/kg/day) given by IP route for 10 days. The significance of difference were tested by using ANOVA followed by Tukey test. Statistical significance depends on the P&lt;0.05.&#x0D; Results: The histological score of cardiac tissue changes showed that a highly increase in mice administrated with trastuzumab when compared to control group while pretreatment with Neuregulin-1 , made a decrease in histological changing score when compared with trastuzumab group. The percentage of immunohistochemical score of stain uptake for TNF-α and IL-1β expression in cardiac tissue shows a highly increase in mice administrated when compared with control group, furthermore, pretreated with neuregulin-1 showed a decrease in the percentage of staining when compared with trastuzumab group.&#x0D; Conclusion: Neuregulin-1 can be used as a cardioprotective in cardiac complications of cancer treatment by its ability to attenuate the cardiotoxic effect of trastuzumab by decreasing in histological changing score and decrease in the percentage of staining when compared with trastuzumab group.

Characterization of Two Novel Bacteriophages Infecting Multidrug-Resistant (MDR) Acinetobacter baumannii and Evaluation of Their Therapeutic Efficacy in Vivo.

Acinetobacter baumannii is emerging as a challenging nosocomial pathogen due to its rapid evolution of antibiotic resistance. We report characterization of two novel bacteriophages, PBAB08 and PBAB25, infecting clinically isolated, multidrug-resistant (MDR) A. baumannii strains. Both phages belonged to Myoviridae of Caudovirales as their morphology observed under an electron microscope. Their genomes were double stranded linear DNAs of 42,312 base pairs and 40,260 base pairs, respectively. The two phages were distinct from known Acinetobacter phages when whole genome sequences were compared. PBAB08 showed a 99% similarity with 57% sequence coverage to phage AB1 and PBAB25 showed a 97% similarity with 78% sequence coverage to phage IME_AB3. BLASTN significant alignment coverage of all other known phages were <30%. Seventy six and seventy genes encoding putative phage proteins were found in the genomes of PBAB08 and PBAB25, respectively. Their genomic organizations and sequence similarities were consistent with the modular theory of phage evolution. Therapeutic efficacy of a phage cocktail containing the two and other phages were evaluated in a mice model with nasal infection of MDR A. baumannii. Mice treated with the phage cocktail showed a 2.3-fold higher survival rate than those untreated in 7 days post infection. In addition, 1/100 reduction of the number of A. baumannii in the lung of the mice treated with the phage cocktail was observed. Also, inflammatory responses of mice which were injected with the phage cocktail by intraperitoneal, intranasal, or oral route was investigated. Increase in serum cytokine was minimal regardless of the injection route. A 20% increase in IgE production was seen in intraperitoneal injection route, but not in other routes. Thus, the cocktail containing the two newly isolated phages could serve as a potential candidate for therapeutic interventions to treat A. baummannii infections.

A systematic review of preclinical studies on therapeutic potential of stem cells or stem cells products in peritoneal fibrosis.

Peritoneal fibrosis remains a serious complication of long-term peritoneal dialysis. Stem cell therapy is an innovative field of scientific investigation with potential for clinical application. Here, we systematically reviewed the studies to determine whether stem cell based therapy could improve the peritoneal fibrosis in experimental models of peritoneal fibrosis. Our systematic search of PubMed, Scopus, Web of Science, and Cochrane Library yield 5219 article. After screening for eligibility, in vivo, experimental, interventional studies using stem cells in animal models of peritoneal fibrosis; 11 articles were included. The studies underwent comprehensive review, quality assessment, and data extraction. Mesenchymal stem cells were the most used type (90.9%) originated either from bone marrow (70%), adipose tissue (20%), or umbilical cord (10%). In 90.9% of studies, stem cells were injected after peritoneal insult and 63.6% of studies used the intraperitoneal injection route. Eight studies met the ≥50% of criteria indicated by ARRIVE recommendation. Information regarding the nature of ethical review permissions, species, strain and gender, dose, route and duration of treatment, was stated by all studies; 81.8% of the studies reported the number of animals in each group. Adverse events were reported in one study. Improvement in histological parameters including attenuation of submesothelial thickness (100%), inflammation (62.5%), angiogenesis (60%), and fibrosis (85.7%) was reported after stem cell therapy. Peritoneal permeability function by assessing the ultrafiltration, glucose transport and solute permeability was improved in all studies. Stem cell treatment resulted in mesothelial recovery in 100% of studies. In preclinical studies, the use of stem cells is associated with improved peritoneal fibrosis. This may provide an important foundation to support future translational clinical research using stem cell therapy to repair the injured peritoneum and modulate immune responses in PD patients.

Assessment of immune response and protection against bacterial coldwater disease induced by a live-attenuated vaccine delivered orally or intraperitoneally to rainbow trout, Oncorhynchus mykiss (Walbaum)

Abstract Flavobacterium psychrophilum is the causative agent of bacterial coldwater disease (BCWD), which has a major global impact on salmonid aquaculture. A candidate live-attenuated F. psychrophilum vaccine strain, CSF259-93B.17 (B17), has been developed and shown to provide significant protection when delivered by intraperitoneal (IP) or immersion routes. This study examined immune response in rainbow trout and protection induced by B17 grown in iron-limited conditions against BCWD, when administered either orally or via injection. B17 was administered via both routes with and without alginate microencapsulation, and comparative protective efficacy was assessed. The microencapsulation method used successfully encapsulated viable B17 bacteria. Fewer microencapsulated B17 cells were available per unit volume of vaccine compared to unencapsulated bacteria. However, protection did not significantly differ for microencapsulated and unencapsulated B17 bacteria regardless of the route of delivery. Despite substantially elevated challenge pressure, both oral and IP delivery achieved significantly better protection than controls, and orally administered B17 achieved protection levels comparable to those achieved by IP immunisation. Serum antibody production response was slower in orally immunised fish, but achieved similar titres to IP immunised fish prior to bacterial challenge. The study indicated that with further optimisation, oral delivery of B17 may form a beneficial component of vaccination strategies, and contribute to the overall management of BCWD at salmonid hatcheries. Statement of relevance F. psychrophilum is the causative agent of bacterial coldwater disease (BCWD), which is a significant impediment to salmonid aquaculture worldwide. In this study, we evaluate the protection and antibody response against BCWD elicited in rainbow trout by immunisation with a live attenuated strain of F. psychrophilum, delivered by oral or intraperitoneal injection routes. In addition, we assess the effect of alginate microencapsulation on the efficacy of this vaccine candidate. We show that it is possible to elicit both an antibody response and measurable reduction in mortality by oral administration of the vaccine, even under elevated pathogenic challenge pressure.

Mir-29a Displays in Vitro and in Vivo Anti-Tumor Activity in Multiple Myeloma

Despite advances in recent therapeutic approaches including targeted therapies, multiple myeloma (MM) remains still incurable necessitating the development of novel treatment strategies. MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate post-transcriptional gene expression and play a critical role in tumor pathogenesis. Tumor suppressor miRNAs are generally down-regulated in cancer cells compared to their normal counterpart, and their enforced expression indeed represents a promising strategy for cancer treatment. In this study, we sought to characterize the role of miR-29a as a tumor suppressor as well as evaluated its therapeutic potential in MM.miR-29a expression levels were found down-regulated in a panel of 5 MM cell lines, 6 newly diagnosed MM patient samples compared to its expression in normal hematopoietic cells collected from 10 normal healthy individuals suggesting that high expression of miR-29a might be involved in MM pathogenesis. We further assessed the functional significance of miR-29a by both gain- and loss-of-function studies. A significant decrease in cell viability (22-32%, p<0.05), along with induction of apoptosis (30-35%, p<0.05) was observed at 48 hrs in MM cell lines, MM.1S and 8226 transfected with miR-29a compared to cells transfected with scrambled miRNA. In contrast, cell lines transfected with miR-29a antagonist prevented the loss of viability in such cells indicating the specificity of miR-29a. At the molecular level, we have identified c-Myc, an important oncogenic transcription factor known to stimulate MM cell proliferation, as a target of miR-29a. Binding site of miR-29a was first identified by computer algorithm and further confirmed by the use of a 3’UTR of c-Myc reporter (luciferase renilla/firefly) constructs containing, miR-29a target site. Moreover, treatment with PRIMA-Met, a small molecule anti-tumor agent in phase I/II clinical trials, significantly increased the expression of miR-29a (2 to 6-fold) and decreased expression of c-Myc in MM cell lines and primay MM patient samples suggesting an important role of miR-29a in inhibiting proliferation of MM cells. On the other hand, overexpression of c-Myc in 8226 and MM.1S cells at least partially reverted the functional effect of miR-29a or PRIMA-1Metsuggesting a specific role of c-Myc in mediating its anti-proliferative activity.To examine therapeutic potential of our studies, we took advantage of novel lipid based delivery method of miRNA. Intratumor delivery of the miR-29a by intraperitoneal injection route against MM xenografts in SCID mice resulted in a significant inhibition of tumor growth (~60%) at 12 days of treatment and prolongation of survival (median survival increased from 22 days to 35 days, p<0.038) compared to the mice receiving scrambled miRNA. Retrieved tumors from treated mice showed efficient increase in miR-29a (5.5-fold, p=0.025), and decrease in c-Myc protein as well as reduced expression of Ki67 and increase of Tunel expression. Similar phenomenon was observed by systematic delivery of miR-29a (by intraveneous injection) in mice with no significant side effects or toxicity in mice.Our study reveals an important role of miR-29a as a tumor suppressor in mediating anti-tumor activities in MM cells by targeting c-Myc. Our findings provide a proof-of-principle that formulated synthetic miR-29a exerts therapeutic activity in preclinical models, and support a framework for development of miR-29a based treatment strategies in MM patients. DisclosuresNo relevant conflicts of interest to declare.

A Bioengineered Peptide that Localizes to and Illuminates Medulloblastoma: A New Tool with Potential for Fluorescence-Guided Surgical Resection.

Tumors of the central nervous system are challenging to treat due to the limited effectiveness and associated toxicities of chemotherapy and radiation therapy. For tumors that can be removed surgically, extent of malignant tissue resection has been shown to correlate with disease progression, recurrence, and survival. Thus, improved technologies for real-time brain tumor imaging are critically needed as tools for guided surgical resection. We previously engineered a novel peptide that binds with high affinity and unique specificity to αVβ3, αVβ5, and α5β1 integrins, which are present on tumor cells, and the vasculature of many cancers, including brain tumors. In the current study, we conjugated this engineered peptide to a near infrared fluorescent dye (Alexa Fluor 680), and used the resulting molecular probe for non-invasive whole body imaging of patient-derived medulloblastoma xenograft tumors implanted in the cerebellum of mice. The engineered peptide exhibited robust targeting and illumination of intracranial medulloblastoma following both intravenous and intraperitoneal injection routes. In contrast, a variant of the engineered peptide containing a scrambled integrin-binding sequence did not localize to brain tumors, demonstrating that tumor-targeting is driven by specific integrin interactions. Ex vivo imaging was used to confirm the presence of tumor and molecular probe localization to the cerebellar region. These results warrant further clinical development of the engineered peptide as a tool for image-guided resection of central nervous system tumors.

Toxicity of endogenous peroxynitrite and effects of puerarin on transplanted retinal pigment epithelial sheets in the subretinal space in mice.

To evaluate the toxicity of endogeneous peroxynitrite on transplanted retinal pigment epithelial (RPE) sheets and the effect of puerarin on their survival in the C57BL/6 mice after RPE sheets have been transplanted into SD rats' subretinal space . C57BL/6 mice eyes were used to culture RPE cells. Ninety-six SD rats were involved in the experiment. They were divided into control (block control), streptozotocin (STZ, negative control), untransplanted RPE (positive control) and transplanted RPE groups respectively. Diabetes was induced in SD rats by intra-peritoneal STZ injection in the latter three groups. Saline was injected into the subretinal space of 24 SD rats in the untransplanted RPE group and primary RPE sheets were injected into the subretinal space of 24 SD rats in the transplanted RPE group. Puerarin (45mg/kg) was administrated into both untransplanted RPE and transplanted RPE groups of diabetic rats through intra-peritoneal injection route after RPE sheets transplantation. At 20, 40, 60 days after surgery, Western blotting analysis, DNA ladder and RT-PCR were used for determining the differences in expression of nitrotyrosine (NT, the foot print of peroxynitrite ), apoptosis and iNOS mRNA in the control, STZ, untransplanted RPE and transplanted RPE groups respectively. HE staining was used for determining the RPE survival in the subretinal space of the transplanted RPE group. Apoptosis and expression of NT and iNOS mRNA were observed in STZ, untransplanted RPE and transplanted RPE groups, but were delayed in untransplanted RPE and transplanted RPE groups in a time-dependent manner compared with control and STZ groups (P<0.01). There were no differences between the two groups (P>0.01). NT, DNA ladder, iNOS mRNA were down-regulated, which were associated with the decrease of expression of peroxynitrite. Numerous pigmented cells emerged and increased in number in the subretinal space during the 60-day observation period after transplantation. On day 20, heavily pigmented cells were visible at the transplant site; On day 40, monolayer and multilayered transplant was visible in the subretinal space; On day 60, heavily pigmented monolayer and multilayered transplants with round apical profile were present along Bruch's membrane. Puerarin increased the 60-day survival of C57BL/6 mice RPE xenografts in the SD rats' subretinal space, which may be related to its direct inhibition of apoptosis of RPE cells and antagnism of damage of peroxynitrite to RPE cells.

Acute physiopathological effects of honeybee ( Apis mellifera) envenoming by subcutaneous route in a mouse model

Bee stings are a health concern in the Americas, where fatal envenomings due to massive attacks by Africanized honeybees have been documented in the last decades. Most studies on the toxic effects of honeybee venom in experimental animals have been performed using the intravenous or intraperitoneal injection routes. The aim of this study was to develop a mouse model that would better resemble a massive honeybee attack by using the subcutaneous (s.c.) route to induce a severe, sublethal systemic envenoming. An array of acute venom effects were characterized, including biochemical, hematological, histological, and inflammatory alterations, after the s.c. injection of 0.5 median lethal dose of venom. Rapid increases in serum alanine (ALT) and aspartate (AST) transaminases, creatinine, urea nitrogen, uric acid, sodium and chloride electrolytes, and creatine kinase (CK) were recorded, indicating damage to liver, kidneys, and skeletal muscle. Also, coagulation disturbances (fibrinogen decrease, and moderate delay in prothrombin and partial thromboplastin times) were demonstrated. Circulating platelet and leukocyte numbers remained unaltered, but a hemoconcentration effect (hematocrit and hemoglobin increase) was observed. This effect might be related to the marked edema induced by the venom. In addition, this inflammatory response included a systemic increase in cytokines (IL-1β, IL-6, TNF-α), together with an elevation of serum malondialdehyde and nitric oxide. The myotoxic effects of venom, melittin, and phospholipase A 2 were demonstrated after injection by s.c. route. No synergistic myotoxicity between melittin and PLA 2 was observed. Moreover, these two components, when injected at equivalent concentrations to those present in venom, induced a lower increase in serum CK than venom, suggesting that other components also contribute to its strong systemic toxicity towards skeletal muscle. The model here presented may be useful in preclinical studies to assess therapeutic antivenoms developed to cope with the problem of massive bee attacks.

Role of Glutamine Depletion in Directing Tissue-specific Nutrient Stress Responses to L-Asparaginase

Role of Glutamine Depletion in Directing Tissue-specific Nutrient Stress Responses to L-Asparaginase

Preclinical assessment of the ability of polyvalent (Crotalinae) and anticoral (Elapidae) antivenoms produced in Costa Rica to neutralize the venoms of North American snakes

Polyvalent (Crotalinae) and anticoral (Elapidae) antivenoms produced by Instituto Clodomiro Picado, Costa Rica, were assessed for their ability to neutralize various toxic activities of the venoms of North American snakes of the genera Crotalus, Agkistrodon and Micrurus, in assays involving preincubation of venom and antivenom. When the intraperitoneal route of injection was utilized, polyvalent (Crotalinae) antivenom was effective in the neutralization of the venoms of Crotalus atrox, Crotalus adamanteus, Crotalus viridis viridis, Crotalus horridus atricaudatus, Agkistrodon contortrix contortrix and Agkistrodon piscivorus piscivorus, whereas the venom of Crotalus scutulatus was not neutralized. When the intravenous route was used, results differed depending on the ‘challenge dose’ of venom employed. Polyvalent antivenom neutralized all venoms when mice were challenged with 2 LD 50s of venom. When 5 LD 50s were used, antivenom neutralized the venoms of C. atrox, C. adamanteus, C. v. viridis and C. h. atricaudatus, being ineffective in the neutralization of C. scutulatus, A. c. contortrix and A. p. piscivorus. Polyvalent antivenom was effective in the neutralization of hemorrhagic and myotoxic activities of all venoms studied. It also neutralized coagulant activity of C. adamanteus venom, whereas most of the venoms were devoid of clotting activity on plasma in vitro. Moreover, it neutralized defibrinating activity of the only three venoms that induced this effect (i.e. C. adamanteus, A. c. contortrix and A. p. piscivorus). Anticoral (Elapidae) antivenom neutralized lethality induced by the venom of Micrurus fulvius, using either the intravenous or the intraperitoneal routes of injection. Moreover, it neutralized myotoxic effect of this venom as well. It is concluded that polyvalent antivenom neutralizes lethality and other activities of most of the crotaline venoms tested. However, since it is ineffective in neutralizing the lethal effect of C. scutulatus venom, it is suggested that a venom containing presynaptically-active neurotoxic phospholipases A 2 related to ‘mojave toxin’ needs to be introduced in the immunizing mixture in order to increase the neutralizing scope of this product in North America. Anticoral antivenom is highly effective in the neutralization of the venom of M. fulvius.

Mechanism of biosynthesis of trimethylamine oxide from choline in the teleost tilapia, Oreochromis niloticus, under freshwater conditions

The mechanism of biosynthesis of trimethylamine oxide (TMAO) from dietary precursors in the teleost tilapia ( Oreochromis niloticus) was investigated. Diets supplemented with quaternary ammonium choline, glycine betaine, carnitine or phosphatidylcholine were administered and significant increases in TMAO levels in the muscle were only observed with choline. [ Methyl- 14C] and [ 1,2- 14C] cholines were given through dietary and intraperitoneal injection routes, but 14C-TMAO was detected only in fish with dietary administration of [ methyl- 14C] choline. Dietary treatment with [ 15N] choline resulted in the formation of [ 15N] TMAO in the muscle. The incorporation of radioactivity into TMAO was also observed both following dietary administration and intraperitoneal injection of [ 14C] trimethylamine (TMA). When choline was introduced into the isolated intestine, marked increases in TMA levels occurred. These increases were significantly suppressed in the presence of penicillin. [ 14C]-TMA derived from [ methyl- 14C] choline was detected in the cavity of the isolated intestine. The introduction of [ 15N] choline into the intestinal cavity resulted in the formation of [ 15N] TMA. TMA mono-oxygenase activities were detected in the liver and kidney. We conclude that tilapia possess the ability to produce TMAO from choline, which is related to intestinal microorganisms and tissue mono-oxygenase under freshwater conditions.

Subcutaneous administration of liposomes: a comparison with the intravenous and intraperitoneal routes of injection

The development of long-circulating liposomes containing lipid derivatives of poly(ethylene glycol) (PEG), termed Stealth® liposomes, has considerably improved the prospects for therapeutic applications of liposomal drug delivery systems. We have examined the pharmacokinetics and biodistribution of long-circulating, as compared to conventional, liposomes after subcutaneous (sc) administration in mice. Results obtained after subcutaneous administration were compared to those obtained after intravenous (iv) and intraperitoneal (ip) administration. Liposomes, following sc administration, appeared intact in the circulation subsequent to moving down the lymph node chains that drain the site of injection. Liposomes containing PEG-distearoylphosphatidylethanolamine (PEG-DSPE) resulted in the highest levels of small (80–90 nm) liposomes in the blood, with up to 30% of in vivo label appearing in the blood at 12 to 24 h post-injection. In the absence PEG-DSPE approx. 4-fold lower levels of liposomes were found in the blood. Small size of the liposomes was critical to their ability to move into the circulation, with liposomes above 110–120 nm not appearing in blood to any significant extent. The presence of PEG-DSPE and cholesterol was important for the in vivo stability of the liposome after sc administration. Although liposome levels were significantly higher in the draining lymph nodes after sc administration, levels associated with other tissues were proportionately reduced relative to the iv and ip routes of administration. Liposomes appeared in blood after ip and sc administration with half-lives of approx. 0.6 and 9 h, respectively, and subsequent to appearing in blood had similar biodistribution, pharmacokinetics and half-lives (20.4 h) to liposomes given by the iv route.

Intrapleural administration of fibres induces mesothelioma in rats in the same relative order of hazard as occurs in man after exposure.

1. The dose-response data for the induction of mesothelioma, in rats, by the intrapleural administration of the fibrous zeolite, erionite, has been compared to the published data for the crocidolite and chrysotile forms of asbestos. Erionite is more than two orders of magnitude more carcinogenic than either of the two forms of asbestos examined. 2. The relative sensitivity of the intrapleural and intraperitoneal routes of injection were also examined. The sensitivity of the intraperitoneal over the intrapleural route of administration was considerably greater for all the forms of asbestos examined but not for erionite. 3. The relationship for different fibres, between the number of fibres required to give animals mesothelioma, at the 50% or 10% observable tumour effect level (OTEL) was examined, and a ranking of relative carcinogenicity was made. 4. This showed that the data derived from the dose responses obtained by the intrapleural administration of fibres to rats ranked the relative carcinogenicity of erionite, crocidolite and chrysotile in accord with the known clinical mesothelioma induction in man after exposure to these fibres. Examination of the carcinogenicity ranking from data derived from intraperitoneal injections of fibres was not in accord with the known clinical mesothelioma induction in man for the various asbestos types examined.

3] Production and characterization of monoclonal idiotypes and anti-idiotypes for small ligands

The procedures described in this chapter have enabled us to identify and characterize monoclonal antibodies and their respective anti-idiotypes. We have developed several different types of immunoassays which afford greater flexibility to the investigator, depending on the type of antibodies desired and the availability of labeled antigens. Use of the intrasplenic injection technique for the final booster immunization prior to the fusion protocol has enabled us to achieve more consistent results than the usual intravenous or intraperitoneal injection routes. Isoelectric focusing of tissue culture supernatant from monoclonal antibody-secreting clones can easily identify possible duplicate clones, and thereby reduces the amount of labor required for extensive characterization of a large number of clones. We have found that these techniques have enabled us to identify "sister clones" or redundancies in our collection of antiligand and anti-idiotype antibodies rapidly and accurately. These various techniques have allowed us to save much time, labor, and money in the search for specific antibodies with desired characteristics.

Distributions of amyloid plaques in four regions of the brains of mice infected with scrapie by intracerebral and intraperitoneal routes of injection.

VM mice were inoculated by intracerebral and intraperitoneal routes with brain homogenates containing the 87V strain of scrapie. The distribution and numbers of plaques were found for the parietal cortex, cingulate cortex, corpus callosum and hippocampus/dentate in coronal sections cut at the level of the thalamus and stained with Masson's trichrome. For the intracerebrally injected animals, the greatest numbers were seen on the side of the brain that had been injected. In the four regions, they were most numerous in the corpus callosum and least numerous in the parietal cortex and hippocampus/dentate. Following an intraperitoneal injection, plaques were absent from the corpus callosum and, in the remaining three regions, they were less numerous than in intracerebrally injected animals, although their relative numbers were similar. The distribution of the plaques was suggestive of an initial passive spread of inoculum following intracerebral injection and a tendency for plaques to be associated with myelinated axon tracts.

Suppression of liver uptake of liposomes by dextran sulfate 500.

The effect of dextran sulfate (DS, 500,000 Mr) and multilamellar vesicles (MLV) as liver blockade agents has been investigated in mice. Intravenous injection of unlabeled MLV prior to radioactive MLV caused moderate reduction in the liver uptake and increased tibia, lung, and spleen uptake. More drastic differences were observed with intraperitoneal injection of DS. When tested in the range of 0-50 mg of DS per kg of body weight, maximal liver blockade occurred at a dose of 50 mg. By using 50 mg of DS per kg, maximal liver blockade occurred at 12 hr after DS injection. The liver blockade was temporary, ending within 48 hr. The intraperitoneal route of injection for DS was more effective for liver blockade than the intravenous route.