Intradermal alpha1-antitrypsin therapy avoids fatal anaphylaxis, prevents type 1 diabetes and reverses hyperglycaemia in the NOD mouse model of the disease - PubMed (original) (raw)

Intradermal alpha1-antitrypsin therapy avoids fatal anaphylaxis, prevents type 1 diabetes and reverses hyperglycaemia in the NOD mouse model of the disease

H Ma et al. Diabetologia. 2010 Oct.

Abstract

Aims/hypothesis: Human alpha1-antitrypsin (hAAT) gene therapy prevents type 1 diabetes in a NOD mouse model of diabetes. However, repeated i.p. injections of hAAT into NOD mice leads to fatal anaphylaxis. The aim of the study was to determine if an alternative route of administration avoids anaphylaxis and allows evaluation of hAAT's potential for diabetes prevention and reversal. We also sought to determine if the addition of granulocyte colony-stimulating factor (G-CSF), augments hAAT's capacity to prevent or reverse disease in the NOD mice.

Methods: To evaluate hAAT pharmacokinetics, serum hAAT levels were monitored in NOD mice receiving a single dose (2 mg) of hAAT by i.p., s.c. or i.d. injection. For studies of type 1 diabetes prevention and reversal, mice received i.d. hAAT (2 mg/mouse/3 days) for 8 or 10 weeks or hAAT and G-CSF (i.p., 6 microg/day) for 6 weeks. Blood glucose determinations, glucose tolerance testing and insulin tolerance tests were performed.

Results: Both i.p. and s.c. injections resulted in fatal anaphylaxis. The i.d. injection avoided anaphylaxis and i.d. injection of hAAT into 11-week-old NOD mice prevented disease (p = 0.005, AAT vs PBS at 40 weeks of age). Treatment of diabetic NOD mice with hAAT or hAAT plus G-CSF provided long-term (at least 100 days) reversal of diabetes in 50% of treated animals. G-CSF did not enhance the reversal rates of hAAT. Glucose tolerance and insulin levels were normalised in mice with hAAT prevention and reversal.

Conclusions/interpretation: Intradermal hAAT prevents and reverses disease in a NOD mouse model of type 1 diabetes without inducing anaphylaxis.

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Figures

Figure 1:

Figure 1:. Pharmacokinetics of hAAT in NOD mice.

Cohorts of 8 week old NOD mice (n=6) were injected with 2 mg of hAAT (Prolastin®). Serum hAAT levels were detected by hAAT specific ELISA. Filled diamond, intraperitoneal injection; open square, subcutaneous injection; filled triangle, intradermal injection; open circle, using an osmotic pump (Alzet® Pump). a, hAAT levels within 60 minutes of administration. b, Long term (7 days) hAAT levels after single administration. c, Area under-the-curve of hAAT levels in the first 7 days (AUC0–7 day) of hAAT levels.

Figure 2:

Figure 2:. Human AAT (hAAT) protein therapy prevents type 1 diabetes development in NOD mice.

Cohorts of 11 week-old female NOD mice were ID injected with hAAT (2 mg/mouse, every 3 days) for 10 weeks (indicated by a gray bar), and/or G-CSF (6 mg/mouse, daily) for 8 weeks (indicated by open bar). Open diamond, hAAT (n=8); filled diamond, hAAT plus G-CSF (n=7); filled triangle, G-CSF alone (n=8); open triangle, saline (n=9). a, Serum levels of hAAT. b, Levels of anti-hAAT antibodies detected by ELISA. c, Life table analysis for diabetes development. At 25 weeks of age, all mice in hAAT and hAAT plus G-CSF treated groups remained diabetes free. At 40 weeks of age (the end of the experiment), 63 % of mice in AAT group were diabetes free (P=0.005, AAT vs PBS).

Figure 3.

Figure 3.. Reversal of type 1 diabetes by hAAT protein therapy.

New onset diabetic mice were treated with hAAT (2mg/mouse), G-CSF (6mg/mouse) alone or saline. Both hAAT and G-CSF treatments were administered for 8 weeks. Each line represents data from an individual animal. a, Blood glucose levels in saline treated group (n=5). b, Blood glucose levels in G-CSF treated group (n=6). c, Blood glucose levels in mice treated with hAAT and successfully reversed from type 1 diabetes (n=4). d. Blood glucose levels in mice treated with hAAT that failed to reverse type 1 diabetes (n=3).

Figure 4.

Figure 4.. Reversal of type 1 diabetes by hAAT and G-CSF therapy.

New onset diabetic mice (n=6) were treated with hAAT (2mg/mouse) and G-CSF (6mg/mouse) for 8 weeks. Each line represents data from an individual animal. a, Blood glucose in long term reversed mice. b, Blood glucose in mice who failed to reverse type 1 diabetes. c, Effect of disease severity on the efficacy of treatments. Starting blood glucose levels (at the onset of diabetes) in different groups are plotted. **, P=0.0044.

Figure 5.

Figure 5.. The effects of hAAT treatment on islet function, insulin resistance and immune system.

a, Intraperitoneal glucose tolerance test (IPGTT). Open diamond, diabetic mice (n=4); filled square, hAAT prevented mice (40 weeks of age, n=4); open triangle, hAAT reversed mice (euglycemic for more that 100 days, n=3); filled circle, normal diabetic free mice (n=4). b, Insulin tolerance test (ITT). c, The effects of hAAT on insulitis. Insulitis was evaluated in AAT prevented mice (40 weeks of age, n=4) and reversed mice (euglycemic for more that 100 days, n=3). The new onset diabetic mice served as control (T1D, n=2).

Figure 6.

Figure 6.. The effect of hAAT on serum BAFF levels.

BAFF levels in hAAT treated group (n=8) and saline treated mice (PBS, n=10) was detected by ELISA at 2 weeks after the treatment. P=0.023 (AAT vs. PBS)

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