Suppressive activity of a macrolide antibiotic, roxithromycin on co-stimulatory molecule expression on mouse splenocytes in vivo

The influence of roxithromycin (RXM) on the expression of co-stimulatory molecules, CD40, CD80 and CD86, was examined in vivo. When BALB/c mice were immunized intraperitoneally with two doses of dinitrophenylated ovalbumin (DNP-OVA) at 1 week intervals, intraperitoneal administration of RXM at 250 microg/kg once a day for 14 days strongly suppressed IgE contents in sera obtained from mice 22 days after the first immunization. In addition, RXM treatment of mice suppressed endogenous IL-4 contents in aqueous spleen extracts, which were enhanced by DNP-OVA immunization. We next examined the influence of RXM on co-stimulatory molecule expression on splenic lymphocytes. RXM treatment of the immunized mice caused suppression of CD40 expression, but this treatment did not affect CD80 and CD86 expression.     

Influences of roxithromycin on cell-mediated immune responses.

We investigated the effects of roxithromycin (RXM), a synthesized macrolide antibiotic on murine cellular immune responses by examining the in vitro proliferative response of lymphocytes, interleukin 1 (IL-1) production and interleukin 2 (IL-2) production. RXM was orally administered to BALB/c mice at a dose of 5 mg/kg once a day for 42 days. Spontaneous blastic activity of lymphocytes prepared from mice administered with RXM for 7 days was higher than those from control mice. The activity peaked at the 14th day, and then decreased gradually to control levels by the 42nd day. Time kinetics of lymphocyte blastogenesis to concanavalin A showed a pattern similar to that observed in spontaneous blastic activity. Oral administration of RXM also influenced cytokine production; short-term (for 14 days) administration of RXM enhanced both IL-1 and IL-2 production but long-term (for 42 days) administration inhibited them.     

Inhibitory action of a macrolide antibiotic,roxithromycin, on co-stimulatory molecule expressions in vitro and in vivo

OBJECTIVE: The influence of a macrolide antibiotic, roxithromycin (RXM), on co-stimulatory molecule expression was examined in vitro and in vivo. MATERIALS AND METHODS: Spleen cells obtained from BALB/c mice 10 days after immunization with 8.0 microg of hemocyanin absorbed to 4.0 mg of aluminum hydroxide were cultured in the presence of 100.0 microg/ml of hemocyanin and various concentrations of RXM. We first examined the influence of RXM on cell activation by examining the proliferative response of cells and cytokine production. We also examined the influence of RXM on co-stimulatory molecule (CD40, CD80 and CD86) expressions on cultured splenic B-lymphocytes induced by in vitro antigenic stimulation using flow cytometry. In the second part of experiments, non-immunized and immunized mice were treated orally with 2.5 mg/kg of RXM once a day for 4 or 8 weeks. Splenic B lymphocytes were obtained from these mice 24 h after antigenic challenge, and co-stimulatory molecule expressions were examined by flow cytometer. RESULTS: Cell activation induced by in vitro antigenic stimulation was suppressed by RXM when cells were cultured in the presence of more than 5.0 microg/ml of the agent. Addition of RXM at a concentration of 5.0 microg/ml into cell cultures also suppressed co-stimulatory molecule (CD40, CD80 and CD86) expressions on splenic B lymphocytes, which was enhanced by antigenic stimulation in vitro. Oral RXM administration for 4 weeks clearly suppressed the enhancement of CD40 and CD86 (but not CD80) expressions on splenic B lymphocytes induced by antigenic stimulation in vivo. This suppressive activity of RXM on co-stimulatory molecule (CD40 and CD86) expressions was further strengthened by the treatment of mice for 8 weeks. Long-term treatment with oral RXM also suppressed CD80 expressions, which was not suppressed by 4-week treatment. CONCLUSION: The present results suggest that RXM exerts its immunomodulating effects through suppression of both cell activation and co-stimulatory molecule expressions induced by antigenic stimulation. These suppressive activities of RXM might contribute, in part, to the therapeutic mode of action of RXM on inflammatory diseases.     

Influence of a macrolide antibiotic, roxithromycin, on mast cell growth and activation in vitro

BACKGROUND: Long-term administration of macrolide antibiotics is recognized to be able to favorably modify the clinical condition of inflammatory diseases, such as diffuse panbronchiolitis and cystic fibrosis. However, the precise mechanisms by which macrolide antibiotics could improve clinical conditions of the patients are not well understood. AIM: The present study was designed to examine the influence of macrolide antibiotics on effector cell functions responsible for inflammation through the choice of roxithromycin (RXM) and mast cell. METHODS: Mast cells were induced by long-term culture of splenocytes from BALB/c mice. RXM was added to the cultures at seeding and then every 4-5 days, when the culture medium was replaced with a fresh one. The influence of RXM on mast cell growth was evaluated by counting the number of cells grown on the 16th day. We also examined the influence of RXM on mast cell activation by examining histamine release and inflammatory cytokine secretion. RESULTS AND CONCLUSION: RXM could not inhibit mast cell growth, even when splenocytes were exposed to 100 microg/ml of RXM throughout the entire culture periods. RXM also could not suppress histamine release from cultured mast cells in response to non-immunological and immunological stimulations. However, RXM could suppress inflammatory cytokine, interleukin-1beta, interleukin-6, granulocyte macrophage-colony stimulating factor and tumor necrosis factor-alpha, secretions induced by concanavalin A stimulation at a concentration of as little as 0.5 microg/ml. These results may suggest that RXM modulated the ability of mast cells to secrete inflammatory cytokines and results in improvement of clinical condition of chronic inflammatory diseases.     

Absence of endogenous interleukin-10 enhanced organ dysfunction and mortality associated to zymosan-induced multiple organ dysfunction syndrome

Experimental evidence suggests that interleukin (IL)-10 plays a pivotal role in generalized inflammation. Here we investigate the effects of IL-10 gene deletion on the acute phase of the multiple organ dysfunction syndrome (MODS) caused by zymosan in the mouse. MODS was induced by zymosan administration (500 mg/kg, suspended in saline solution, i.p.) in IL-10 wild-type and knockout mice; sham groups were treated with vehicle. Mice were sacrificed 18 h after zymosan or saline administration. In another set of experiments, animals were monitored for 12 days to assess systemic toxicity and survival rate. Mice lacking IL-10 displayed increased peritoneal exudate volume and leukocytes. Also, we observed a significant increase in myeloperoxidase activity and lipid peroxidation in ileum and lung tissues, as well as augmented levels of TNF-alpha, IL-1beta and nitrogen-derived species in the plasma. With regard to organ injury, absence of IL-10 enhanced the renal, hepatocellular and pancreatic dysfunction caused by zymosan administration. All of these parameters significantly influenced the systemic toxicity and the overall survival at 12 days, which was significantly lower in IL-10 knockout mice. Therefore, this study demonstrates that the absence of endogenous IL-10 enhances the MODS induced by zymosan in mice.     

The challenge of continuous exogenous glucocorticoid administration in mice

Background

Chronic administration of exogenous glucocorticoids is often required in experimental research. We compared the efficacy and reliability of three different methods of continuous glucocorticoid administration in mice.

Materials and methods

Male CD1 Swiss White mice aged 7-9 weeks received corticosterone (CS) or carrier by either subcutaneous (s.c.) injection (n = 15), s.c. implantation of micro-osmotic pumps (n = 20) or s.c. implantation of slow-release pellets (n = 20). Serial blood samples were taken for the measurement of plasma CS and osteocalcin (OC). Bone structural parameters were analysed by micro-computed tomography (μ-CT) in animals treated via slow-release pellets for 4 weeks.

Results

Injection of CS (10 mg/kg) resulted in peak plasma CS levels of up to 2600 μg/L after 1 h, with levels returning to baseline within 4 h post-injection. Micro-osmotic pumps failed to consistently alter plasma CS levels and had variable effects on plasma OC levels. Implantation of 10 mg CS pellets induced hypercorticosteronemia within 24 h but levels returned to baseline within 7 days. Plasma OC levels fell rapidly on day 1 and remained suppressed until day 7. Weekly replacement of pellets maintained elevated plasma CS and suppressed plasma OC concentrations, and resulted in significant bone loss at the tibia and spine after 28 days.

Conclusion

Once-weekly s.c. implantation of slow-release pellets to mice appears to result in relatively consistent plasma CS and OC levels with significant biological effects. However, at least in our hands, no method delivered CS at a constant rate and variability in plasma CS levels was pronounced.     

Suppressive activity of a macrolide antibiotic, roxithromycin, on pro-inflammatory cytokine production in vitro and in vivo

This study was designed to examine the influence of a macrolide antibiotic, roxithromycin (RXM), on the production of pro-inflammatory cytokines, interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha. In the first experiments, we examined the effect of RXM on in vitro cytokine production from lipopolysaccharide (LPS)-stimulated human peripheral blood monocytes. The monocytes were cultured in the presence of various doses of the agent. After 24 h, the culture supernatants were obtained and assayed for IL-1beta and TNF-alpha contents by enzyme-linked immunosorbent assay. RXM suppressed the in vitro production of IL-1beta and TNF-alpha in response to LPS stimulation. This was dose dependent and first noted at a concentration of as little as 0.05 microg/ml, which is much lower than therapeutic blood levels. In the second part of the experiments, we examined the influence of RXM on the appearance of IL-1beta and TNF-alpha in mouse lung extract induced by LPS inhalation. RXM was administered orally into BALB/c mice at a single dose of 2.5 mg/kg once a day for 5-12 weeks. These mice were then instilled with LPS into the trachea and examined for the presence of cytokines in aqueous lung extracts. Pretreatment of mice with RXM for 5 weeks did not influence of the appearance of both IL-1beta and TNF-alpha in aqueous lung extracts. However, pretreatment for more than 7 weeks dramatically suppressed the cytokine appearance in the extracts.     

Influence of carnitine supplementation on muscle substrate and carnitine metabolism during exercise

We examined 1) the effect of L-carnitine supplementation on free fatty acid (FFA) utilization during exercise and 2) exercise-induced alterations in plasma levels and skeletal muscle exchange of carnitine. Seven moderately trained human male subjects serving as their own controls participated in two bicycle exercise sessions (120 min, 50% of VO2max). The second exercise was preceded by 5 days of oral carnitine supplementation (CS; 5 g daily). Despite a doubling of plasma carnitine levels, with CS, there were no effects on exercise-induced changes in arterial levels and turnover of FFA, the relation between leg FFA inflow and FFA uptake, or the leg exchange of other substrates. Heart rate during exercise after CS decreased 7-8%, but O2 uptake was unchanged. Exercise before CS induced a fall from 33.4 +/- 1.6 to 30.8 +/- 1.0 (SE) mumol/l in free plasma carnitine despite a release (2.5 +/- 0.9 mumol/min) from the leg. Simultaneously, acylated plasma carnitine rose from 5.0 +/- 1.0 to 14.2 +/- 1.4 mumol/l, with no evidence of leg release. Consequently, total plasma carnitine increased. We concluded that in healthy subjects CS does not influence muscle substrate utilization either at rest or during prolonged exercise and that free carnitine released from muscle during exercise is presumably acylated in the liver and released to plasma.     

Ascorbic acid decreases heat shock protein 70 and plasma corticosterone response in broilers (Gallus gallus domesticus) subjected to cyclic heat stress

It is known that ascorbic acid (AA) supplementation can ameliorate the chicken's responses to heat stress. The influence of AA on heart heat shock protein 70 (hsp70) and plasma corticosterone (CS) was evaluated in young male broiler chickens fed either no AA (N-AA) or 500 mg AA /kg (AA) and exposed to cyclic high temperatures (21 to 30 to 21 degrees C) over a 3.5 h period on three consecutive days. Dietary AA supplementation elevated plasma AA and maintained it at high levels after heating, but in N-AA birds, only heat elevated plasma AA. In N-AA fed chickens, plasma CS was elevated and was further increased by heat stress as compared with AA-fed birds. Heart hsp70 expression was greater in N-AA-fed chickens compared to AA-fed chickens, and heat stress further elevated hsp70 in both N-AA- and AA-fed birds. The hsp70 increase after heat was two-fold greater in N-AA- vs. AA-fed birds. Plasma CS and heart hsp70 were positively correlated, plasma AA and heart hsp70 were negatively correlated, and plasma CS and AA were negatively correlated. It was concluded that chickens experience a less severe stress response after exposure to high temperatures when they are provided dietary AA.     

Factors modifying the effect of diazepam on plasma corticosterone levels in rats

We have examined factors that alter the effect of diazepam (DZ) on plasma corticosterone (CS) in rats. DZ had a biphasic effect on plasma CS levels: CS decreased with doses below 5 mg/kg and increased with higher doses. Peak response occurred 90 minutes post injection in both sexes. Plasma DZ levels were significantly higher in females than in males and peak at 10 and 30 minutes post injection in males and females, respectively. There was also a sex difference in the pattern of DZ metabolites. An acute stressor (30 minutes of immobilization) did not affect plasma CS levels in rats injected with a 5 mg/kg dose of DZ. Prenatally stressed animals did not differ in basal CS levels or in their response to 5 mg/kg of DZ compared to prenatally non-stressed animals. These two groups of animals also did not differ in plasma levels of DZ or of its metabolites. By contrast, the 5 mg/kg dose of DZ had no effect on plasma testosterone levels in control animals, but increased it in prenatally stressed animals. Furthermore, compared to non-stressed controls, prenatally stressed animals had lower baseline plasma testosterone levels. These results indicate that the effect of DZ on plasma CS is influenced by endogenous as well as exogenous factors and that these effects vary with the particular biochemical parameter under examination.     

Insulin-like growth factor I prevents corticosteroid-induced diaphragm muscle atrophy in emphysematous hamsters

The aim of this study was to evaluate whether recombinant human insulin-like growth factor I (rhIGF-I) could attenuate or prevent diaphragm (DIA) fiber atrophy with corticosteroid (CS) administration to emphysematous (EMP) hamsters. DIA muscle IGF-I responses to CS administration with and without exogenous rhIGF-I administration were evaluated. Three groups were studied: 1) EMP; 2) EMP + triamcinolone (T; 0.4 mg.kg-1.day-1 im); and 3) EMP + T + IGF-I (600 microg/day by constant infusion). After 4 wk, the DIA was analyzed histochemically and biochemically (IGF-I mRNA levels by RT-PCR and endogenous and exogenous IGF-I peptide levels immunochemically). Body weights of EMP-T progressively decreased, while those of EMP and EMP-T-IGF-I remained stable despite similarly reduced food intake in both T groups. DIA weight was reduced with T but preserved with rhIGF-I infusion. DIA fiber proportions were similar among the groups. The cross-sectional areas of types I, IIa, and IIx fibers were reduced (17 to 31%) with T administration but unchanged with rhIGF-I infusion. DIA IGF-I mRNA levels were similar across all groups. By contrast, the endogenous DIA IGF-I levels were reduced (41%) in the EMP-T-IGF-I animals. Total DIA IGF-I levels (endogenous + exogenous) were still significantly reduced. IGF-I immunoreactivity confirmed this reduction in all DIA fibers. We conclude that DIA fiber atrophy with T was completely prevented by exogenous rhIGF-I administration. This effect was likely mediated by the pharmacological influences of exogenously administered rhIGF-I. We speculate that this results from increased bioavailability of free IGF-I to react with muscle receptors. Reduced endogenous IGF-I levels in the DIA likely reflect a negative-feedback influence. These results may have important clinical implications for treatment options to offset the adverse effects of CS on the respiratory muscles in patients with chronic lung disorders.     

Modulation of human neuroblastoma transplanted into nude mice by endogenous opioid systems

The role of endogenous opioid systems (endogenous opioids and opioid receptors) in human cancer was explored using an opioid antagonist paradigm and neuroblastoma cells (SK-N-MC) transplanted into nude mice. Mice inoculated with 2.5 X 10(6) neuroblastoma cells received daily injections of either 0.1 or 10 mg/kg naltrexone (=0.1 and 10 NTX groups) which blocked the opioid receptor for 6-8 hr/day or the entire 24 hr/day, respectively, or sterile water. The latency for appearance of a measurable tumor (5 mm diameter) in the 0.1 NTX group was 27% longer than controls (11 days), and the first death in this group occurred 33% later than controls (day 27). Mice inoculated with tumor cells in the 10 NTX group had an acceleration (18%) in the latency of tumor appearance and, 2 weeks after cell inoculation, 70% of the mice in this group had tumors, in contrast to 10% of the controls. At the termination of the experiment (day 45), only 33% of the 10 NTX group were alive, in contrast to 90% of the controls. Receptor binding assays using DAGO, DADLE, or EKC revealed specific saturable binding only for DADLE and EKC. NTX administration resulted in a 148-186% increase in density for both binding sites, but no changes in binding affinity. Measures of opioid levels showed that tumor tissue levels of both beta-endorphin and methionine-enkephalin were elevated 2.5 to 6.5 fold from control values in both NTX groups, whereas plasma beta-endorphin was subnormal by 4 to 6 fold. These results indicate that endogenous opioid systems regulate human neuro-oncogenesis, with opioids being active inhibitors of growth. Opioid antagonists up-regulate receptors and increase tissue levels of endogenous opioids and, under conditions in which the opioid antagonist is short-acting (e.g., 0.1 NTX), can have an exaggerated antitumor effect during the interval when the antagonist is no longer present.     

Opioidergic control of gonadotropin secretion in the female rabbit: divergent effects of morphine on secretion of follicle-stimulating hormone and luteinizing hormone

The nature of secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) was followed in female rabbits on a daily basis from age 36 to 60 days by sequential 5-min blood sampling over 1- to 2-h periods each day. Both LH and FSH were found to be secreted in a pulsatile manner. The mean LH pulse amplitude over the 25 days was 0.95 +/- 0.32 ng/mL and for FSH it was 10.15 +/- 1.11 ng/mL. Mean plasma LH levels were significantly increased from 1.46 +/- 0.08 ng/mL in 36 to 42-day-old rabbits to 1.89 +/- 0.12 ng/mL in 43 to 50-day-old rabbits and remained elevated from 50 to 60 days. FSH levels during the same periods also rose significantly from 14.93 +/- 0.79 to 19.57 +/- 2.05 ng/mL. To examine the influence of endogenous opioid peptides on the release of LH and FSH in 36 to 60-day-old female rabbits, morphine sulfate at 0.2, 0.5, 2.0, and 5.0 mg/kg was administered subcutaneously after 30 min baseline sampling, and blood was taken for another 60-120 min. Morphine at all doses and at all ages inhibited the amplitude and frequency of LH pulses but had no effect on FSH secretion. To determine whether the effects of morphine on LH secretion could be reversed with naloxone, females aged 82-114 days were used. Naloxone administered 1 h after morphine reversed the inhibitory effects of morphine, whereas the simultaneous administration of naloxone with morphine had variable effects but seemed to delay the LH increase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Extinction of a conditioned response in rainbow trout selected for high or low responsiveness to stress

Two lines of rainbow trout (Oncorhynchus mykiss) that exhibit divergent endocrine responsiveness to stressors also display disparate behavioral traits. To investigate whether the high-responding (HR) and low-responding (LR) fish also differ in cognitive function, the rate of extinction of a conditioned response was compared between the two lines. Groups of HR and LR fish were exposed to a paired conditioned stimulus (CS; water off) and unconditioned stimulus (US; confinement stressor). After exposure to 18 CS-US pairings, at least 70% of individuals of both lines acquired a conditioned response (CR) manifested as an elevation of blood cortisol levels on presentation of the CS only. Post-conditioning, the fish were tested by presentation of the CS at weekly intervals, for 4 weeks, with no further reinforcement, and the extinction of the CR in the two lines was compared. The decline in mean plasma cortisol levels after exposure to the CS over successive tests suggested that the CR was retained for a shorter period among the HR (<14 days) than LR fish (<21 days). The frequency of individuals within each line whose plasma cortisol levels indicated a stress response when exposed to the CS was significantly greater among the LR than HR fish at 14 and 21 days with no HR fish falling into this category at 21 days. At 28 days post-conditioning, there were no HR fish and only three LR fish were categorized as "stressed". These results suggest that there are differences in cognitive function between the two lines. Possible mechanisms underlying these differences are discussed.     

Effects of postnatal treatment with naloxone on plasma gonadotropin,prolactin, testosterone and testicular functions in male rats

Opioid peptides are implicated in the control of gonadotropin and prolactin secretion. The role of opioid antagonist naloxone and its effects on plasma gonadotropin, prolactin, testosterone levels and testicular hyaluronidase, acid phosphatase, [³H]uridine and thymidine incorporation, RNA, DNA and protein concentrations were evaluated in rats after administration of naloxone beginning day 1 through 21 and autopsied on 45, 60 and 90 days of age. Plasma gonadotropin and testosterone levels were significantly elevated after naloxone treatment. Testicular hyaluronidase and acid phosphatase activity increased till 60 days post treatment and declined thereafter. Concentrations of RNA and protein did not change significantly but the concentration of DNA declined at 45 and 60 days of age. These results suggest that endogenous opioid peptides exert regulatory influence on gonadotropin secretion which in turn control the testicular function in the male rat.     

The effect of elevated blood cortisol levels on the extinction of a conditioned stress response in rainbow trout

Following previously published observations that a conditioned response (CR) was lost more quickly by rainbow trout (Oncorhynchus mykiss) exhibiting a high responsiveness to stressors than by low responding individuals this study was designed to investigate the effects of exogenous cortisol on the retention of a CR in unselected rainbow trout. Fish held in isolation were conditioned over a 10-day period by pairing an innocuous signal (conditioned stimulus, CS: a water jet played on the surface of the tank water) with a mild stressor (unconditioned stimulus, US: 30 min of confinement). This resulted in a brief elevation of plasma cortisol levels (the CR) when the fish was exposed to the CS only. The effect of exogenous cortisol on the retention of the CR was evaluated by comparing the performance of fish that received cortisol-containing slow-release intraperitoneal implants, with fish receiving vehicle-only implants. Retention of the CR was assessed at intervals up to 35 days after conditioning ceased. The CR was considered to be evident when 30 min following presentation of the CS, mean plasma cortisol levels were significantly higher in conditioned than untrained fish. On day 1 both cortisol-implanted and vehicle-implanted conditioned fish exhibited a CR. However, from day 5 onwards the CR was observed only in the vehicle-implanted and conditioned group. This finding indicates that administration of cortisol accelerated the extinction of the CR in the cortisol-implanted fish, suggesting that elevated plasma cortisol levels can impair memory processes in rainbow trout.     

In vivo stimulatory effect of Cordyceps sinensis mycelium and its fractions on reproductive functions in male mouse

Cordyceps sinensis (CS), an Ascomycetes fungus parasitic to Lepidoptera larvae, has been traditionally used as nutritious food for the enhancement on sexual performance and the restitution of impairment in sexual function in Chinese society. We have previously demonstrated the stimulatory effect of CS and its fractions on steroidogenesis both on primary mouse Leydig cells and MA-10 mouse Leydig tumor cells. In the present studies, we determined the in vivo effects of CS and its fractions on steroidogenesis in mouse. Different concentrations of CS and CS fractions (0.02 and 0.2 mg/g body weight) were fed to immature or mature mice from 1 to 7 days. The plasma levels of testosterone were evaluated by radioimmunoassay. The weights of reproductive organs were also determined. Results illustrated that CS significantly induced plasma testosterone levels both in immature and mature mice in 3 and/or 7 days treatment (p < 0.05). F2 and F3 at 0.02 and/or 0.2 mg/g body weight for different feeding duration could also significantly stimulated plasma testosterone levels both in immature and mature mice (p < 0.05). In general, CS, F2 and F3 didn't have considerable effect on the weights of reproductive organs. Taken together, these studies illustrate that CS and its fractions significantly stimulated in vivo mouse testosterone production.     

Effects of epidermal growth factor on serum zinc and plasma prostaglandin E2 levels of mice with pressure sores

This work was undertaken to study the effects of systemic application of EGF on the plasma PGE2 and serum zinc levels of mice with pressure sores. The pressure sores were made on the hind limb of the mice by bilateral sciatic nerve neurotomy. Mice were injected IP, 1 μg EGF daily for ten days, begining on the 21st day after operation. EGF administration increased the serum zinc and plasma PGE2 levels compared with controls. These results indicate that EGF can be effective on wound healing by elevating the serum zinc and plasma PGE2 concentrations together with other physiological roles in cellular events.     

Combined exposure to cigarette smoke and nontypeable Haemophilus influenzae drives development of a COPD phenotype in mice

Background

Cigarette smoke (CS) is the major etiologic factor of chronic obstructive pulmonary disease (COPD). CS-exposed mice develop emphysema and mild pulmonary inflammation but no airway obstruction, which is also a prominent feature of COPD. Therefore, CS may interact with other factors, particularly respiratory infections, in the pathogenesis of airway remodeling in COPD.

Methods

C57BL/6 mice were exposed to CS for 2 h a day, 5 days a week for 8 weeks. Mice were also exposed to heat-killed non-typeable H. influenzae (HK-NTHi) on days 7 and 21. One day after the last exposure to CS, mice were sacrificed and lung inflammation and mechanics, emphysematous changes, and goblet cell metaplasia were assessed. Mice exposed to CS or HK-NTHi alone or room air served as controls. To determine the susceptibility to viral infections, we also challenged these mice with rhinovirus (RV).

Results

Unlike mice exposed to CS or HK-NTHi alone, animals exposed to CS/HK-NTHi developed emphysema, lung inflammation and goblet cell metaplasia in both large and small airways. CS/HK-NTHi-exposed mice also expressed increased levels of mucin genes and cytokines compared to mice in other groups. CS/HK-NTHi-exposed mice infected with RV demonstrated increased viral persistence, sustained neutrophilia, and further increments in mucin gene and chemokine expression compared to other groups.

Conclusions

These findings indicate that in addition to CS, bacteria may also contribute to development of COPD, particularly changes in airways. Mice exposed to CS/HK-NTHi are also more susceptible to subsequent viral infection than mice exposed to either CS or HK-NTHi alone.     

Administration of semisynthetic human insulin by a spray injector

The use of Jet injection in insulin administration pointed out the question whether this route could affect insulin absorption and plasma insulin profiles. To compare plasma insulin profiles following an administration of an identical insulin dose by jet injection or by conventional subcutaneous route (syringe with needle) 8 healthy subjects (age 24-28 yrs., non obese) were given at 09.00 h of two different days 200 mU/kg/BW of human semisynthetic regular insulin (Novo Actarapid) alternatively subcutaneously by a syringe with needle or transcutaneously by jet injection (DG 77 - Sicim - Gorizia). Before insulin administration and then 15, 30, 60, 90, 120 and 180 minutes after, blood samples were drawn for plasma insulin and C-peptide determination. Higher plasma insulin levels after administration by jet were found at 15' and 30' minutes (62,58 +/- 6,31 v.s. 36,94 +/- 3,31 microunits/ml at 15' and 76,51 +/- 9,60 v.s. 51,65 +/- 9,95 at 30', p less than 0,01 and p less than 0,005, paired Student t test). No difference could be observed for the other times. C-peptide was found to fall to undetectable values, confirming the nearly total suppression of endogenous insulin production. It is concluded that total regular insulin absorption does not differ after transcutaneous jet injection or administration by syringe with needle, but in the first case it is faster. This last finding should be considered in planning insulin treatment schedules.