RESEARCH COMPENDIUM

November, 2006

Table of Contents

I. Introduction

II. History and Biochemistry of various insulin therapies

III. A description of intensive IV insulin therapy

IV. Summary of Published Studies on IV insulin

V. Current Studies

a. Clinical Effects of IV insulin on Cognitive Function in Patients with Diabetes Mellitus

b. Effects of IV insulin in Treatment of Diabetic Patients with Nephropathy

c. Effects of IV insulin in the Treatment of Diabetic Patients with Heart Disease

d. Effects of IV insulin on Diabetic Patients with Neuropathy

e. Effects of IV insulin on Progressive Retinopathy in Patients with Diabetes Mellitus

f. Effects of IV insulin in Diabetic Patients with nonhealing wounds

g. Biochemistry IV insulin on circulating blood markers

h. Effects of Effects of IV insulin on Hepatic Metabolism in a Diabetic Rat Model.

i. Phase II Cognitive Study – Correlation of Enhanced Memory Function in IV insulin Treated Diabetic Patients with Functional Magnetic Resonance Imaging

j. Phase II Neuropathy Study – Correlation of Neuropathy Improvement in Diabetic Patients undergoing IV insulin with Quantitative Neurologic Measures

k. Effects of IV insulin in Patients with Brittle Types 1 and 2 Diabetes

I. Introduction

Diabetes is one of the most serious challenges to healthcare worldwide and is projected to affect 239 million people by the year 2010, a doubling in the prevalence since 1994 (1). Diabetes is the most common cause of progressive and disabling neuropathy in the world. Diabetes leads to great morbidity and mortality and results in a significant economic impact on society, both in terms of healthcare resources and lost productivity in the workplace (2,3). Diabetic neuropathy is the most common form of neuropathy in the developed countries of the world, and is responsible for 50-75% of non-traumatic amputations (2,4).

The precise mechanism of how metabolic abnormalities initiate and promote the development secondary complications is not currently known. However, these early changes in metabolic function precede the development of irreversible structural damage to the cells and nerves.

We believe that the metabolic environment plays an early and central role in the development and progression secondary complications. Furthermore, failure to rectify the poor metabolism has a prolonged impact on the progressive pathologic changes. The development of neurovascular complications of diabetes begins with an underlying genetic predisposition which, when acted upon by initiating events such as overfeeding or smoking, results in inflammatory changes that may precede hyperglycemia (5). Inflammation and hyperglycemia unleash a cascade of events that effect cellular proteins, gene expression, and cell-surface receptor expression in the endothelium, ultimately resulting in progressive pathologic changes and subsequent vascular complications.

II. History and Biochemistry of Insulin

Normally, insulin is secreted during digestion. As with many hormones, insulin is secreted in discrete bursts over a one hour period. When hormones are not delivered in bursts there appears to be down regulation in cells particularly in the liver. This down regulation is known to decrease the hormones’ metabolic effectiveness. Down-regulation at the cellular level may partially explain the decreased action of steady-state levels, while discrete bursts of hormones may allow recovery of receptor affinity or receptor numbers. For maintenance of insulin-dependent enzymes essential for glucose metabolism (e.g. hepatic glucokinase, phosphofructokinase, and pyruvate kinase), require a defined insulin level (200-500 µU/ml in the portal vein) in combination with high glucose levels (bimolecular signal). In non-diabetic subjects, when insulin first reaches the liver, 50% of the insulin is removed, this along with the evolutionary fact that the pancreas is directly connected to the liver by the portal vein, implies that the liver is the principal metabolic target organ of both insulin and the carbohydrates produced during digestion. As stated by Becco, Aoki “The insulin retained by the hepatocytes may itself be essential for the long-term effects of insulin on hepatic glucose metabolism as well as growth and de novo enzyme synthesis. Following oral glucose intake, the liver accounts for an equal or greater portion of total net glucose uptake compared to the periphery. Insulin exerts pivotal control of glucose levels through its ability to regulate HGP directly or indirectly. The traditional subcutaneous (S.C.) insulin administration regimens used by diabetic patients a) fails to capture the pulsatile nature of natural insulin secretion and b) does not reach high enough insulin concentrations at the hepatocyte level (e.g., 10 U regular insulin injected S.C. produce a peak systemic circulation concentration of 30-40 µU/ml and an even lower portal vein concentration of 15-20 µU/ml).”

As Gecco and Aoki went on to report; “A relative deficiency of insulin at the hepatocyte level leads to an impaired capacity to process incoming dietary glucose. With the liver functioning as the target organ of the pancreas, it can be concluded that the primary purpose of giving insulin to the diabetic patient should not be to control blood glucose level (“control theory”) but rather the normalization of hepatic metabolism. Furthermore, these same hepatic enzymes are found in all glucose-utilizing bodily systems, suggesting a synchronous effect by insulin and glucose.”

It is well understood that Type I diabetics and some type II diabetics have a resting Respiratory Quotient (RQ) of .7 to .8. This RQ reflects the bodies’ problem processing glucose as the primary source of energy. The bodies’ inability to utilize glucose in the presence of insulin suggests a some missing key to metabolism. When these patients with low RQs are given high doses of insulin and large amounts of carbohydrates, their RQs change from .7-.8 to .98-1.05. This change in RQ indicates that they have switched from fatty acid metabolism to glucose metabolism. The switch to glucose metabolism signifies a fundamental metabolic shift which leads to significant improvement in secondary complications associated with diabetes.

III. Protocol

IV insulin which mimics the bodies natural rhythms promotes the normalization of carbohydrate metabolism in diabetic patients. IV insulin effects multiple organs, especially muscle, retina, liver, kidney, and nerve endings. The process involves the administration of high-dose insulin similar to those found in the portal circulation of normal humans. The process is monitored by frequent glucose levels and respiratory quotients (RQ). RQ is measured by a metabolic cart which determines the ratio VCO₂/ VO₂. This ratio is specific for the fuel used at any one time by the body. The glucose levels are monitored to keep glucose levels appropriate, and the RQ determines the need to readjust the infusion.

IV insulin, which mimics natural patterns, increases the respiratory quotient from levels around 0.7 to levels greater than 0.9. This reflects the underlying physiology of the treatment, confirming the conversion from fat metabolism, typical in the diabetic patient, to a normal metabolic state utilizing carbohydrate as the primary fuel consumed. In order to most accurately mimic normal metabolism and therefore maximize results, the total amount of insulin given as well as the total amount of consumed glucose is altered with each treatment.

Summary of Clinical trials to Date:

Effect on Glycemic Control

Aoki et al Long-term intermittent intravenous insulin therapy and type 1 diabetes mellitus. Lancet. 1993, 342: 515-517.

This study examined 20 “brittle” diabetics (defined as patients with wide glucose swings and frequent hypoglycemic episodes) treated with the treatment protocol. This was a prospective study with patients serving as their own historic controls. All patients had been on intensive insulin therapy (four shots daily) for at least one year prior to entrance into the study. The results of this study were as follows:

A significant decline in HbA1c from the baseline of 8.5% to 7.0% at the end of the observation period (p < 0.0003)
A decline in the frequency of major hypoglycemic events from 3.0 to 0.1 per month (p < 0.001)
A decline in the frequency of minor hypoglycemic events from 13.0 to 2.4 per month (p < 0.001)

. Effect on Hypertension

Aoki et al., Effect of chronic intravenous insulin therapy on antihypertensive medication requirements in IDDM subjects with hypertension and nephropathy. Diabetes Care (1995): 1260-1265.

This is a prospective, randomized, crossover study, examine antihypertensive medication requirements in 26 hypertensive insulin dependent diabetic patients. These patients were randomized into treatment and control groups. All patients were stabilized prior to the study on four shots of insulin daily and antihypertensive medications (ACE inhibitors, calcium channel blockers, loop diuretics, and alpha two agonists). Following three months in either the treatment or control group, the patients were crossed over into the other group. Total antihypertensive medication requirements were then tabulated.

Antihypertensive dosage requirements decreased significantly (46%, p<0.0001) and linearly over time during the treatment phase, while remaining stable in the control group. Following the crossover, the previously treated patients (now controls) returned to their baseline antihypertensive needs within the subsequent three months.

.Effect of Diabetic Nephropathy

Aoki et al., Effect of intensive insulin therapy on progression of overt diabetic nephropathy in patients with IDDM. Endocrine Practice (1999) 5: 174-178

This is a multicenter, retrospective, longitudinal study, involving 31 patients with type 1 diabetes mellitus and overt diabetic nephropathy. All patients were on intensive (four daily shots) insulin therapy and weekly pulsed IV insulin. All patients were on ACE therapy and aggressive antihypertensive regimens. All patients were followed with creatinine clearance measurements.

Patients were followed for an average of 37 months. Creatinine clearance remained essentially unchanged during this period. These observations suggested that pulsed IV insulin could successfully stabilize renal function in patients with diabetic nephropathy.

Dailey, et al. Effects of pulsatile intravenous insulin therapy (PIVIT) on the progression of diabetic nephropathy. Metabolism (2000) 49: 1491-1495.

The Dailey study is a multiinstitutional prospective, randomized, controlled study evaluating the effect of pulsed IV insulin in patients with diabetic nephropathy. This study included diabetic centers at Mayo Clinic, Scripps Clinic, Joslin Diabetes Center, University of Maryland, University of Arizona, and Temple.

49 patients with type 1diabetes and chronic kidney disease were randomized into treatment and control groups in an 18 month study. Treatment group patients had a statistically significant improvement in renal function as compared to the control group.

Effect on Diabetic Autonomic Neuropathy

Aoki et al.. Effect of intensive insulin therapy on abnormal circadian blood pressure pattern in patients with type 1 diabetes mellitus. Online J Curr Clin Trials (1995) 199.

This is a randomized, controlled clinical study evaluating the abnormal circadian blood pressure pattern in insulin dependent diabetic (IDDM) patients and its response to MAT. 74 IDDM patients were randomized to a treatment group or a control group. 24 hour blood pressure monitoring was performed monthly along with HbAIC levels. All study patients were evaluated weekly by investigators and all were on four shots of insulin daily prior to and during the study.

Following three months of, the night/day systolic BP ratios decreased from 0.97 to 0.94 and increased from 0.95 to 0.98 in the control group (p = 0.0224). The night/day diastolic BP ratio decreased from 0.93 to 0.90 in the treatment group and increased from 0.91 to 0.94 in the control group (p=0.0037. This improvement of an autonomic neural pathway is consistent with a small series reported separately of IDDM patients with severe uncontrollable postural hypotension improving after two months of MAT (Aoki et al., Am J Med (1995) 99: 683-684.

Background of Expanded Clinical Studies

Following the publication in 2000 of the independent multiinstitutional study (Mayo Clinic, Scripps Clinic, Harvard’s Joslin Diabetes Center, University of Maryland, University of Arizona, and Temple) demonstrating pulsed IV insulin effectiveness in stabilizing renal disease (see above), many related questions naturally followed:

1. Could these findings be expanded to other diabetic complications?

2. Is the effect of pulsed IV insulin on patients with type 2 diabetes analogous to its demonstrated effect on patients with type 1 diabetes?

3. Could this treatment be reproduced in a consistent fashion in multiple clinical settings in order to develop a powerful tool for the treatment of patients with severe diabetic complications?

Advanced Diabetes Treatment Centers, LLC was established to answer the above questions. The mission of ADTC is to identify board certified endocrinologists with a special interest in the care of the diabetic patient, and to establish centers with those physicians to further the understanding of the effect of IV insulin treatment on the significant complications of diabetes.

By teaming up with Dr. Betty Tuller of Florida Atlantic University, the vast resources of the Florida University system has facilitated interspecialty participation and cooperation in furthering the study of pulsed IV insulin . In addition, cooperative studies involving outside universities and physicians (Bascom Palmer Eye Institute at the University of Miami (retinopathy study), East Virginia Medical School (neuropathy study), and Johns Hopkins’ Wilmer Ophthalmological Institute (retinopathy study)) have expanded the ability to study and to evaluate pulsed IV insulin.

STUDY DESIGNS

All human trials involving IV insulin are approved by a federally sanctioned institutional review board. All studies are performed in a prospective, controlled fashion. Endpoint parameters are scored in a coded blind fashion. Patients in the treatment group undergo IV insulin treatments according to the protocol as outlined above. Two treatment sessions (consisting of three treatments per session) are performed initially, followed by weekly treatments for the duration of the study. The control group undergoes weekly evaluations by the clinic nurse, including vital signs, blood pressure checks, and interim history checks. Any adjustments to their daily medical regimens as dictated by glucose levels or blood pressure are done during these weekly visits.(this is not valid, the patient is then referred to their endocrinologists to make changes to their medical regimen)

Testing specific to any particular study is performed prior to the initiation of IV insulin therapy and at intervals as specified in the study protocol.

All patients must meet entrance criteria for pulsed IV insulin prior to qualifying for any clinical study

Current Entry Criteria for IV insulin treatment based on the above clinical studies and independent clinical observations, the following criteria have been developed for patients to qualify for IV insulin treatment. All patients must be under the care of an endocrinologist, be compliant with their medical regimens, and be willing to undergo the weekly IV insulin treatment.

Patient has HgbA1c >8.0 in spite of 2 or more injections of short acting and/or intermediate acting insulin daily

Patient has hypoglycemia unawareness. This is a major and unique indication for IV insulin treatment

Patient has labile diabetes (sugars ranging from <60 to >300 more than 2 days a week) in spite of multiple insulin injections (>2) or the use of an insulin pump

Patient has significant proteinuria (>300 mg/24 hrs) in spite of ACE inhibitors and/or ARB’s

Patient’s creatinine clearance is less than 60ml/min and declining at greater than 1 ml/month on ACE/ARB’s and insulin therapy

Patient’s blood pressure is uncontrolled (>130/80) on more than 3 BP drugs with a HgbA1c greater than 8.0% on more than 2 insulin shots daily

Patient has progressive, severe diabetic peripheral neuropathy

Patient has orthostatic hypotension due to autonomic neuropathy of advanced diabetes

Patient has advanced gut neuropathy with gastroparesis or diabetic diarrhea

Patient has a non-healing diabetic foot ulcer (no or minimal evidence of healing over 2 months) in the absence of surgically remediable LE ischemia, gangrene and/or osteomyelitis

Patient has progressive retinopathy.

COMPLIANCE REVIEW:

Patient is willing and able to devote up to 6 hours a week over a minimum of six months to IV insulin treatment

Patient is currently compliant with his/her diabetes treatment regimen.

IF PATIENT HAS ANY OF ABOVE INDICATIONS AND COMPLIANCE REVIEW IS FAVORABLE, PATIENT IS A LIKELY CANDIDATE FOR TREATMENT

Current Clinical Studies of the Center for Complex Systems and Brain Sciences, Florida Atlantic University

A. Clinical Effects of IV INSULIN on Cognitive Function in Patients with Diabetes Mellitus

Introduction

Cognitive impairment is common in patients with diabetes mellitus, especially for memory and executive function (see [1] for review). Moreover, patients with type 2 diabetes, especially women over age 65, not only show lower levels of cognitive function but also increased rates of cognitive decline relative to age- and sex-matched cohorts [2]. The growing awareness of cognitive impairments in diabetes is especially intriguing given the well documented presence of insulin receptors in brain tissue that are selectively expressed in areas of the brain associated with memory functions and deteriorate in Alzheimer’s disease [3]. Furthermore, Watson and Craft [4] demonstrated temporary improvement of memory functions following the administration of intravenous insulin in a subtype of Alzheimer’s patients with insulin resistance in these receptors.

Protocol

This is a prospective, controlled, single-blind study examining cognitive function in patients undergoing treatment. Patients receive cognitive testing before beginning treatment and at three month intervals thereafter. Cognitive testing includes several modified Wechsler III memory tasks (immediate and delayed recall of stories and word lists). In addition, serial laboratory studies which may include CBC, chemistry profile, TSH, B12, folate, hbA1C, and urine protein. These results are compared with an age, sex, and disease matched control group who also undergo the cognitive testing every three months, and the laboratory studies. Entry criteria include diabetic patients, aged 21 – 80, without other primary causes of cognitive impairment (brain tumor, previous neurosurgery, memory impairing medications). These patients must meet one or more criteria for treatment (see Criteria for treatment above)

Patients undergo treatment as noted in previous protocols weekly over a period of 6-12 months with renewals for successive 6-month periods. Control group patients undergo weekly visits with the clinic nurse with vital sign and glucose checks as described above. All testing is recorded for later off-line analysis by an independent researcher blind to the patient’s grouping.

Endpoints and Statistics:

Endpoints include measurements of changes in short and long term memory abilities as indicated by the cognitive tests.

Statistical evaluation uses repeated measures ANOVA with one grouping factor (treatment vs. control group) and multiple regressions (the latter to evaluate effects of age, sex, and duration of diabetes).

References:

[1] Stewart R, Liolitsa D: Type 2 diabetes mellitus, cognitive impairment and dementia. Diabetic Medicine 16: 93-112 (1999). PMID 10229302

[2] Gregg EW, Yaffe K, Cauley JA, Rolka DB, Blackwell TL, Narayan V, Cummings SR: Is diabetes associated with cognitive impairment and cognitive decline among older women? Arch Intern Med 160: 174-180 (2000). PMID 10647755

[3] Steen E, Terry BM, Rivera EJ, Cannon JL, Nelly TR, Tavares R, Xu XJ, Wands JR, dela Monte SM. Impaired insulin and insulin-like growth factor expression and signaling mechanisms in Alzheimer's disease--is this type 3 diabetes? J. Alzheimers Dis 7: 63-80 (2005). PMID 15750215[4] Watson GS, Craft S. Modulation of memory by insulin and glucose: neuropsychological observations in Alzheimer’s disease. Eur. J Pharmacol 490: 97-113 (2004). PMID 15094077

B. clinical effects of Insulin Therapy in the Treatment of Diabetic Patients with Nephropathy

Introduction

Diabetic nephropathy is a progressive complication leading to end stage kidney disease with anemia, hypertension and eventually dialysis. A multicenter trial of IV Insulin in type 1 diabetics showed a slowing of the progression over a control group using maximal conventional therapy (1). Diabetic nephropathy develops in 20-50% of type 2 diabetics and is one of the most common causes of end stage renal disease (2). Proteinuria occurs early in the disease and continues despite maximal treatment although the rate may slow. This study is to determine the effect of treatment on both types of diabetes in patients with nephropathy.

Protocol

Entry criteria are patients (age 21-80) who have diabetes with serum creatinine over 1.2 mg/dl, minimal to severe proteinuria, and no evidence of other kidney disease (renal obstruction, renal tumors). This is a prospective, randomized, single blinded control study. . Treatment groups will undergo weekly treatments for 12 months with self-renewing 12 month periods. Laboratory studies to be done before insulin therapy may include CBC, chemistry profile, HbA1C, TSH, lipid profile, C- peptide, and urine protein studies. Research labs to be assessed every 6 months are fructosamine, aldosterone, brain natiuretic protein, TGF-beta, endothelin 1, fibrinogen, creatinine clearance and urine protein excretion. The study will be evaluated every 6 months statistically and compared with the control group by the study parameters noted.

Endpoints and Statistics

Endpoints: serum creatinine, creatinine clearance, degree of proteinuria, blood pressure

Statistics: ANOVA

Blood: 1 purple, 2 red tops: Urine: 24 hour overnight collection for protein and creatinine

References:

1. Daily GE, Boden GH, Creech RH, Johnson DG, Gleason RE, Kennedy FP, Weinrauch LA, Weir M, D’Lia JA, Effects of Pulsatile Intravenous Insulin Therapy on the Progression of Diabetic Nephropathy, Metabolism 49:1491-95, 2000. PMID: 11092517

2. American diabetes Association, Position Statement: Nephropathy, Diabetes Care, 26:S94-98,2003.

C. clinical effects of IV Insulin Therapy in the Treatment of Diabetic Patients with Heart disease

Introduction

. Altered glucose metabolism in the heart makes fatty acids a primary energy source (1). Fatty acids require increased oxygen utilization for energy production while producing decreased contractility (2). Improved glucose metabolism has been shown to improve intimal thickness (3). Glucose metabolism is preferred by the heart muscle but impaired in diabetic patients by insulin resistance and decreased post-prandial glucose uptake. This study is designed to test the effect of treatment on cardiac function in diabetic patients with significant cardiac disease (4).

Protocol

Individuals selected (ages 21 and older) will have diabetes with impaired cardiac function. For inclusion into the study, the patient must exhibit one of the following:

1. Impaired cardiac function as noted by NYHA (class 2 or class 3)

2. Abnormal ejection fraction as determined by echocardiography (30-50% of predicted normal)

3. Documented coronary artery disease

4. Diffuse cardiomyopathy

Exclusion criteria include significant renal disease, right sided heart failure, valvular heart disease, or significant pulmonary disease.

Patients selected will undergo treatment for 12 months with carotid ultrasound to determine intimal thickness, echocardiography, and questionnaires before treatment and every 3 months. The study has renewals of successive 6-month periods. Laboratory studies may include CBC, chemistry profile, TSH, hemoglobin A1C, aldosterone, fructosamine, endothelin 1, homocysteine, free fatty acids, and lipids.

Endpoints and Statistics

Endpoints: intimal thickness, ejection fraction, wall motion, and cardiac questionnaire. Laboratory levels: aldosterone, endothelin 1, homocysteine, free fatty acids

Statistics: ANOVA

Blood: two redtops

References:

1. Parley D and Pepin EC, Ischemic Heart Disease: Metabolic Approaches to Management, Clin Cardiology 27:439-41, 2004.

2. Hutter JD, Piper HM, Spieckerman PG, Effects of Fatty Acid Oxidation on Efficiency of Energy Production in Rat Heart, Am J Physio 24: H723-28, 1985.

3. Esposito K, Giugliano D, Nappo F, Marfellen R Regression of Carotid Atheroschlerosis By Control of Postprandial Hyperglycemia in type 2 Diabetes, Circulation 110:214-19, 2004.

4. Shah A, Shannon R, Insulin Resistance in Dilated Cardiomyopathy, Rev Cardiovasc Med 4 (supp 6) S50-57, 2003.

D. Clincal Effects of IV INSULIN THEREPY on Diabetic Neuropathy

Introduction

Diabetic neuropathy (DN) is a progressive complication causing serious problems in 25%-40% of diabetics. Significant complications produce painful peripheral dysthesias, loss of sensation, and gastroparesis. DN may affect the peripheral motor and sensory nerves in addition to the autonomic nervous system (1-3). Treatment strategies for patients with DN have generally concentrated on pain relief, without addressing the underlying pathophysiology of the disease (4). Anecdotal reports from patients undergoing this treatment for other complications suggest that this treatment may show efficacy in patients with DN. This study is designed to compare patients with DN who receive treatment with a control population.

Protocol

Patients to be selected may have type 1 or 2 diabetes on oral agents or insulin and neuropathy primarily related to diabetes, preferably age 21-80, and the study will be 12 months in duration with renewals of successive 6 month periods. The laboratory studies that may be performed include CBC, chemistry profile, TSH, A1C and urine protein. Other research blood markers to be determined may be drawn baseline and every 6 months thereafter. Questionnaires on the neuropathy will be completed every 6 months as well.

Endpoints and Statistics

Endpoints: modified Michigan neuropathy questionnaire or Norfolk Quality of Life Neuropathy questionnaire.

Statistics: ANOVA

Blood: 2 red tops

References:

1. Tesfaye S, Chaturvedi N, Eaton SEM, Ward JD, Manes C, Ionescu-Tirgoviste C, witte DR, Fuller JH, Vascular Risk factors and Diabetic Neuropathy N Engl J Med 352:341-50, 2005.

2. Neuropathy Trust, Diabetic Neuropathy: Prevalence, www.neurocentre.com.

3. Potter PJ, Maryniak O, Yamorski R, Jones IC, Incidence of Peripheral Neuropathy in the Contralateral Limb of Persons with Unilateral Amputation due to Diabetes, Journal of Rehabilitation Research and Development 35:335-39, 1998.

4 Goldstein DJ, Lu Y, Detke MJ, Lee TC, Iyengan , Duloxetine versus Placebo in Patients with Painful Diabetic Neuropathy, Pain 116:109-18, 2005.

E. clincal Effects of IV INSULIN THEREPY on Progressive Retinopathy with Diabetes Mellitus (Protocol developed in conjunction with Scott Cousins, MD, former Professor of Ophthalmology, Bascom Palmer Eye Institute, current Professor of Ophthalmology, Duke University Medical Center and Ingrid Zimmer-Galler, MD, Department of Ophthalmology, the Johns Hopkins Hospital Wilmer Eye Institute)

Diabetic retinopathy is one of the leading causes of blindness in the world. Signs of retinopathy are detected in almost 100% of type 1 diabetic patients who have had their disease for at least 20 years and almost 100% of type 2 diabetic patients with the similar duration of disease (1). Histopathologic findings range from microaneurysms and cotton wool spots to more ominous neovascularization. The latter process, known as proliferative diabetic retinopathy, can progress to total blindness if untreated. The biochemical mechanisms responsible for PDR have been extensively studied, and appear to be multifactorial. Associated findings include abnormalities of vasoactive peptides such as vascular endothelial growth factor (VEGF), pigment epithelium derived factor (PEDF), and insulin-like growth factor (ILF-1), lipids, oxidative pathways, enzymatic pathways, such as protein kinase, and carbohydrate metabolism (1-4). Whether these (and other) factors are interrelated or have a common underlying defect is unknown. The common endpoint, however, is vascular leakage with neovascularization. Current therapeutic regimens based on these biochemical abnormalities have to date been unsuccessful in stemming the progression of proliferative diabetic retinopathy. Current treatment strategies emphasize glycemic and blood pressure control, with laser photocoagulation and vitrectomy for advanced cases (5).

Early retinal disease in diabetic patients may take the form of diabetic macular edema (DME). This is observed in 20% to 25% of both type 1 and type 2 diabetic patients. The pathophysiology of DME involves the leakage of plasma from small vessels in the macula. Resorption of this fluid followed by hard exudate formation can lead to severe impairment of central vision (6).

Anecdotal evidence from ophthalmologic institutions (Houston Eye Institute, Shands at University of Florida, Bascom Palmer Eye Institute) suggests that this treatment arrests the progression of retinal disease in patients with proliferative diabetic retinopathy. The mechanism of this effect is unknown, but may be related to reversal of retinal ischemia or downregulation of vasoactive peptides by restoration of hepatic metabolism.

Protocol

This study is designed as a prospective, controlled, single blinded evaluation of IV Insulin in the role of diabetic retinopathy. The patients entered into the study will be from two distinct sources. First, in conjunction with a national eye imaging company, patients with known type 1 or type 2 diabetes will be evaluated for retinal disease. This evaluation will consist of mydriatic fundus photography in diabetic patients not having had recent ophthalmologic evaluation (period greater than 12 months). The fundus photographs will be read by an observer under the auspices of the Wilmer Ophthalmologic Institute at Johns Hopkins Hospital. Three classifications of patients will be evaluated in this study:

I Patients with non high risk proliferative diabetic retinopathy

II Patients with severe non proliferative diabetic retinopathy

III Patients with non clinically significant diabetic macular edema

Patients who are diagnosed as one of these three classifications will be offered entrance into the study. Study patients will be matched for age, sex, and disease severity into a treatment and control group. All study patients will be evaluated in conjunction with an ophthalmologist and have thorough ophthalmologic evaluation prior to entrance. This evaluation will include clinical examination, fluoroscein angiography, and optical coherence tomography. Treatment group patients will undergo weekly sessions as per protocol above. Control group patients will have weekly clinic visits to maximize glycemic and hypertensive control. All patients will repeat their fundus photography at three month intervals, with ophthalmologic evaluation as above every six months, or more often if requested by the ophthalmologist.

Additional entry criteria in this study includes patients aged 21 to 80 years of age who have no other serious form of eye disease. Patients must be under good glycemic and hypertensive control. Exclusion criteria include patients with severe high risk proliferative retinopathy.

Endpoints and Statistics

Endpoints: Serial fundus photography will be interpreted by treatment blinded grader and scored as to stabilization, progression, or improvement of appearance. In addition, episodes of bleeding and necessity for intervention will be evaluated in all groups. Changes in lab values, may include VEGF, C peptide, Endothelin-I, and hsCRP for anaylsis.

Statistics: ANOVA of laboratory tests; blinded analysis of treatment success by retina specialist

Blood: 2 red tops

References:

1. Frank, R.N. Diabetic retinopathy. NEJM 350 (1) 48-58 (2004).PMID 14702427

2. Caldwell, RB et al. Vascular endothelial growth factor and diabetic retinopathy: pathophysiological mechanisms and treatment perspectives. Diabetes Metab Res Rev. 19(6): 442-55 (2003).PMID 14648803

3. Singleton, JR et al. Microvascular complications of impaired glucose tolerance. Diabetes 52(12): 2867-73 (2003) PMID 14633845

4. Kumar, MA et al. The role of lipids in the development of diabetic microvascular complications: implications for therapy. Am J cardiovasc Drugs 3(5): 325-338 (2003). PMID 14728067

5. Sarraf, JA and Fong, D. Preventing diabetic retinopathy through control of systemic factors. Curr Opin Ophthalmol 14(6) 389-94 (2003). PMID 14615645

6. Fong DS, Aeillo L, Gardner N, King GL, Blankenship G, Cavellerado J, Ferris F Klein R, Diabetic Retinopathy:Position Statement, Diabetes Care, 26:S99-102, 2003. PMID 12502630

F. BIOCHEMISTRY OF IV INSULIN-Clinical Effect of treatment on Circulating Risk Markers of Vascular and Metabolic Complications

Introduction

Insulin produces vasodilatory, anti inflammatory and anti thrombotic effects (1-4). . However the effects of IV Insulin on circulating risk factors for vascular and metabolic disease is unknown. This study is used to evaluate circulating risk markers of vascular and metabolic disease compared to a matched control group.

Protocol

Patients selected have diabetes mellitus, 21 years of age and older, and are treated with oral agents and/or insulin. The study is for a minimum of 12 months and may continue for 1-2 years if a significant difference is shown following the initial 6 months. Blood markers will be determined every 6 months for the first year, and every 6 months after that. They may include the following: BNP, fructosamine, PAI-1, fibrinogen, homocysteine, endothelin 1, aldosterone, VCAM, ICAM, IGF-1, TGF-beta, TNF-alpha, hs-CRP, and IL-6). Other markers may be added to the study although the total blood volume drawn will not change. The results are compared to an age and fructosamine matched control group.

Endpoints; Changes in markers

Statistics: ANOVA

Blood: 2 purple, 2 red, 2 blue tops

References:

1. Katakam PVG, Tulbert CD, Snipes JA, Erdos B, Miller AW, Busija DW, Impaired Insulin-induced Vasodilation in Small Coronary Arteries of Zucker Obese rats is Mediated by Reactive Oxygen Species, AJP-Heart 288:854-60, 2005.

2. Chakraborty K Sinha AK, The Role of Insulin as an Antithrombotic Humoral Factor, BioEssays 26:91-98, 2003.

3. Elias AN, Eng S, Homocysteine Concentrations in Patients with Diabetes Mellitus-Relationship to Microvascular and Macrovascular Disease, Diabetes, Obesity and Metabolism 7:117-21, 2005.

4. Patiag D, Qu X, Gray S, Idris I, Wilkes M, Seale JP, Donnely R, Possible Interactions between Angiotensin II and Insulin:Effects on Glucose and Lipid Metabolism in vivi and in vitro, Journal of Endocrinology 167: 525-31, 2000.

G. clincal Effects of insulinTherapy on Hepatic Metabolism in a Diabetic Rat Model

Introduction.

The treatment depends crucially on exploiting the normal pulsatile secretion of insulin in non-diabetic individuals. By intravenously delivering insulin in a natural fashion, IV Insulin increases the respiratory quotient from levels around 0.7 to levels greater than 0.9. This reflects the underlying physiology of the activation treatment, confirming the conversion from fat metabolism, typical in the diabetic patient, to a normal metabolic state utilizing carbohydrate as the primary fuel consumed. Nevertheless, we do not yet have definitive evidence that peripheral administration of pulsed insulin maintains its pulsatile form at the portal vein. In this work, we use an animal model, diabetic (Zucker) rats, to explore directly whether the form of insulin delivery in a peripheral vein is maintained at the entry to the liver. In future work, we will sacrifice the rat and analyze tissue from the liver to assay the amount of insulin uptake and the level of liver enzymes. In addition, levels of compounds (IRS-1, IRS-2, and AKT) that are integral components of insulin signaling pathways [1] and indicate the status of arterial function [2] will be analyzed from several types of tissue from the diabetic rats.

Protocol.

The first phase of the proposed work investigates whether pulsed insulin in a peripheral vein results in pulsed levels of insulin at the portal vein.

A diabetic rat model (Zucker rats) will be used to investigate both the timing and amplitude of insulin measured at the portal vein. When initially administered via a peripheral vein in the hindlimb. To this end, a cannula will be inserted in a hind limb of the rat and another in the portal vein. Baseline levels of insulin will be recorded and, if necessary, maintained by low-dosage subcutaneous insulin drip. A bolus of insulin will be injected into the peripheral vein and a blood sample taken at the portal vein, every 15 sec for a period of 3 minutes. The bolus will be varied in both amplitude and duration of administration so that the appropriate delivery parameters for the rat can be discovered.

In essence, this protocol will 1) establish a dose response curve for pulsed insulin delivery; 2) determine whether the insulin that gets to the liver maintains its pulsatile character; and 3) determine the amount of insulin that is preserved at the level of the portal vein.

Endpoints and statistics

Amplitudes and durations of measurable insulin levels above baseline will be measured at the portal vein. Standard dose response curves will be determined for insulin levels and durations administered in the periphery and measured at the portal vein. Uptake in the liver will be assessed using 125 I labeled insulin and using radiomicrography on homogenized liver tissue, in order to determine percent total insulin uptake. ANOVA will be used to evaluate percent total insulin uptake relative to baseline and to the bolus injected at the periphery.

References

[1] Taniguchi CM, Ueki K, Kahn R: Complementary roles of IRS-1 and IRS –2 in the hepatic regulation of metabolism. J Clin Invest 115: 718-727 (2005).

[2] Okon EB, Chung AW, Rauniyar P, Padilla E, Tejerina T, McManus BM, Luo H, van Breman C: Compromised arterial function in human Type 2

H. Phase II Neuropathy Study – Correlation of Neuropathy Improvement in Diabetic Patients undergoing insulin therapy with Quantitative Neurologic Measurements

Introduction

Initial studies done (part D) showed a 51% improvement in pain relief and a 42% improvement in sensation. The study of this significant effect of MAT has been amplified by adding quantitative and qualitative testing of peripheral nerve function. Additionally, a more inclusive questionnaire has been substituted to better elucidate the areas of improvement and autonomic testing has been added to evaluate the effect of treatment on this form of neuropathy, a risk factor for sudden death (1, 2).

Protocol

Patients with diabetic neuropathy, ages 21 or older, who are on oral agents and/or insulin are eligible for this study. Nerve conduction studies (Neurometrix Corp) are done initially to evaluate for diabetic neuropathy (3) and to exclude those with other forms of neuropathy. Those that are included in the study are then tested with quantitative testing for hot/cold and vibratory sensation (MEDOC Corp) (4) and may include an autonomic dysfunction using cardiac beat-to-beat variation (5) (Ansar Corp). The Norfolk neuropathy quality of life questionnaire is used and continued monthly while the objective nerve testing is done every 6 months. Treatment group patients will be compared with an age, sex, and disease matched control group.

Endpoints: 1) Changes in nerve conduction, sensory testing and/or autonomic testing. 2) Changes on questionnaire

Statistics; ANOVA

Bibliography

1. Vinik AI, Mehrabyan A, Diabetic Neuropathies, Med Clin N Am 88:947-999, 2004.

2. Koury CB, ADA Releases Diabetic Neuropathies Statement, Diabetic Microvascular Complications Today, 2:18-20, 2005.

3. Vinik AI, Emley MS, Megerian JT, Gozani SN, Median and Ulnar Nerve Conduction Measurements with the Symptoms of Diabetic Peripheral Neuropathy Using the NC-Stat System, Diabetes Technology and Therapeuticss, 6; 816-24, 2004.

4. Zinman LH, Bril V, Perkins BA, Cooling Detection Thresholds in the Assessment of Diabetic Sensory Polyneuropathy, Diabetes Care 27:1674-79, 2004.

5. Curtis BM, O Keefe JH, Autonomic Tone as a Cardiovascular Risk Factor: The Dangers of Chronic Fight or Flight, Mayo Clin Proc 77: 45-54, 2002.

I. clincal Effectiveness of IV INSULIN on “Brittle” Diabetes AND GLUCOSE CONTROL

Introduction

Diabetes can produce wide swings in blood glucoses with erratic control even under optimal conditions. Reasons for this include insulin resistance, erratic insulin secretion, and counterregulatory hormones.(1,2) This can occur despite minimal changes in overall diabetic control as evidenced by hemoglobin A1c levels. This study was instituted to evaluate the effect of IV Insulin on improving diabetic control in these circumstances.

Protocol

Diabetics over age 21 referred for erratic control and elevated hemoglobin A1c despite optimal therapy are placed on treatment weekly. Weekly glucose diaries and glucose monitor recorded glucoses are reviewed after a 30 day baseline before treatment to determine if the treatment improves control. The number of glucoses above 300 mg/dl and below 70 mg/dl as well as mean glucose levels are determined. Hemoglobin A1c and/or fructosamine levels are determined every 3 months. Treatment continues for 6 months and then continues if improvement is noted.

Endpoints; Hemoglobin A1c, fructosamine, mean and SD of glucose levels, number of glucoses above and below specific levels

Statistics: ANOVA

Blood: none

Bibliography

1.Quinones MJ, Nicholas SB, Lyon CJ, Insulin Resistance and The Endothelium, Current Diabetes Reports 5:246-53, 2005.

2.Parrish R, Petersen KF, Mitochondrial Dysfunction and Type 2 Diabetes, Current Diabetes Reports 5:177-183,2005.

Bibliography

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2. Baron AD: Insulin and the vasculature-old actors, new roles. J Invest Med 1196;44:406-412

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4. Bjorntorp P, Schlersten T, Gottfries A: Effects of glucose infusion on adipose tissue lipogenesis in man. Acta Med Scand 1968; 183:565-571.

5. Bomboy JD, Lewis SB, Lacy WW, Sinclair-Smith CC, Liljenquist JE: Transient stimulatory effect of sustained hyperglucagonemia on splanchic glucose production in normal and diabetic man. Diabetes 1977; 26:177-181.

6. Bratush-Marrain PR, Komjati M, Waldhausl WK: Effacacy of pulsatile versus continuous insulin administration on hepatic glucose production and glucose utilization in type I diabetic humans. Diabetes 1986; 35:922-926.

7. Cahill GF Jr, Ashmore J Renold AE, Hastings AB: Blood glucose and the liver. Am J Med 1959; 26:264-282.

8. Cruickshank EWH, Startup CW:The effect of insulin on the respiratory quotient, Oxygen consumption, sugar utilization and glycogen synthesis in the normal mammalian heart hyper- and hypoglycemia. J Physiol 1933; 77:365-385.

9. Ewing DJ, Campbell IM, Clark BF: The natural history of diabetic autonomic neuropathy. QJ mEd 1980; 49-95-108.

10. Green DA, Sima AAF, Feldman EL, Stevens MJ: Diabetic neuropathy screening and management. Contemp Intern Med 1993; 5:79-91.

11. Hostetter TH: Diabetic nephropathy. N Engel J Med 1985; 312:642-643.

12. Ikeda T, Matsabaro T, Sato J Sakamoto M: Circadian blood pressurevariation in diabetic patients with autonomic neuropathy. J Hypertens 1193; 11:581-587.

13. Lehman EO, Gosling RG, Sinksen PH: Arterial wall compliance in diabetes. Diabet Med 1192; 9:114-119.

14. Linger C Favre L,Assal J-Ph:Twenty-four blood pressure and heart profiles of diabetic patients with abnormal cardiovascular reflexes. Diabetic Med 199;8:420-427.

15. Polonsky KS: The B cell in diabetes: from molecular genetics to clinical research Diabtes 1195; 44:705-717.

16. Polonsky KS, Givens BD, Canter EV: Twenty-four-hour profiles and patterns of insulin secretion in normal and obese subjects. J Clin Invest 1988; 81:442-448.

17. Vinik AI, Holland MT, Le Beau JM, Luissi FJ: Diabetic Neuropathies. Diabetes Care 1192; 15:1926-1975.

18. Weinhouse S Regulation of glucokinase in liver. In: Horecker B.L., Stadtman E.R. (eds). Current topics in cell research. New York: Academic Press 1976, 1-50.