wound care for doctors PPP

wound care for doctors PPP

Wound Care (Skin Ulcers)

Chronic skin wounds affect an estimated 6.5 million people in the United States, with treatment costs accounting for up to $25 billion per year. Populations at risk for chronic wounds include the elderly and those with comorbidities such as diabetes and obesity. If not treated properly, chronic wounds can lead to severe local infection, sepsis, tissue or limb amputation, or death.

Types of Skin Ulcer Wounds

Diabetic Ulcer Venous Statis Ulcer Decubitus Ulcer

Risk Factors of Skin Ulcers

You’re more likely to get skin ulcers if you have certain risk factors. These include:

Pregnancy. During pregnancy, hormonal changes and increased blood volume may cause leg vein problems. Cigarette smoking. Tobacco smoke hardens your arteries and disrupts proper blood flow. Limited mobility. Being bedridden, paralyzed, or using a wheelchair puts your skin under constant pressure. Leg injuries and arthritis can limit your movement. Increasing age. Age is linked to atherosclerosis and venous insufficiency. High blood pressure. Hypertension, or high blood pressure, damages the arteries and disrupts blood flow. High blood cholesterol. High cholesterol increases narrowing and oxidative stress in the arteries, which disrupts blood flow. Diabetes. Ulcers form due to a combination of factors, such as lack of feeling in the foot, poor circulation, foot deformities, irritation (such as friction or pressure), and trauma, as well as duration of diabetes Obesity. Obesity raises your risk for diabetes, atherosclerosis, and increased pressure in your leg veins. History of blood clots. If you’re prone to blood clots, you’re more likely to have blood flow issues.

Complications from skin ulcers

If left untreated, a skin ulcer may become infected. This can prolong the healing process. The infection can also spread to deeper tissue, bones, joints, and blood.

Diabetic Foot Ulcer

What is a Diabetic Foot Ulcer?

A diabetic foot ulcer is an open sore or wound that occurs in approximately 15 percent of patients with diabetes and is commonly located on the bottom of the foot. Of those who develop a foot ulcer, 6 percent will be hospitalized due to infection or other ulcer-related complication.

Causes

Diabetes is the leading cause of non-traumatic lower extremity amputations in the United States, and approximately 14-24 percent of patients with diabetes who develop a foot ulcer will require an amputation. Foot ulceration precedes 85 percent of diabetes-related amputations. Research has shown, however, that development of a foot ulcer is preventable. Anyone who has diabetes can develop a foot ulcer. Native Americans, African Americans, Hispanics, and older men are more likely to develop ulcers. People who use insulin are at higher risk of developing a foot ulcer, as are patients with diabetes-related kidney, eye, and heart disease. Being overweight and using alcohol and tobacco also play a role in the development of foot ulcers. Ulcers form due to a combination of factors, such as lack of feeling in the foot, poor circulation, foot deformities, irritation (such as friction or pressure), and trauma, as well as duration of diabetes. Patients who have diabetes for many years can develop neuropathy, a reduced or complete lack of ability to feel pain in the feet due to nerve damage caused by elevated blood glucose levels over time. The nerve damage often can occur without pain, and one may not even be aware of the problem. Your podiatrist can test feet for neuropathy with a simple, painless tool called a monofilament. Vascular disease can complicate a foot ulcer, reducing the body's ability to heal and increasing the risk for an infection. Elevations in blood glucose can reduce the body's ability to fight off a potential infection and also slow healing.

Treatment

Symptoms

Because many people who develop foot ulcers have lost the ability to feel pain, pain is not a common symptom. Many times, the first thing you may notice is some drainage on your socks. Redness and swelling may also be associated with the ulceration and, if it has progressed significantly, odor may be present.

Decubitus Ulcer (Sacral Pressure Ulcer)

Pressure ulcers, also known as bedsores or decubitus ulcers, are skin injuries that develop most commonly on bony areas of the body. A sacral pressure ulcer is one of the most common types of these injuries. The sacral region is the area of the lower back near the spine. “Sacral” refers to the sacrum which is the tailbone, or the triangular pelvic bone where most people rest their weight while sitting.

Causes

Pressure ulcers (also known as pressure sores or bedsores) are injuries to the skin and underlying tissue, primarily caused by prolonged pressure on the skin. They can happen to anyone, but usually affect people confined to bed or who sit in a chair or wheelchair for long periods of time. Constant pressure to any part of your body can lessen the blood flow to the tissue. Friction occurs when the skin rubs against clothing or bedding making fragile skin more vulnerable to injury, especially when it is moist. Shear occurs when two surfaces move in the opposite direction. For example, when a bed is elevated at the head, you can slide down in the bed. As the tailbone moves down, the kin over the bone might stay in place – essentially pulling in the opposite direction. The main cause of bedsores among seniors is nursing home neglect. Elders are at a higher risk of bedsores if they cannot easily move on their own. Bedsores typically develop when someone cannot reposition their body over a long period of time. Without movement, the skin loses blood flow and eventually decays.

Stages of a Decubitus (Pressure) Ulcer

Not all pressure ulcers are serious or life-threatening when they are diagnosed. Doctors classify pressure ulcers by stages based on the depth and severity of the wound. The four stages of pressure ulcers are: Stage 1: The skin is red and irritated, but still intact. The area of the sore is painful. It no longer has healthy blood supply and left untreated can progress to an open wound. A Stage 1 pressure ulcer can form after just a few hours of unrelieved pressure on the skin of a vulnerable patient. That is why preventative treatment is vitally important. Stage 2: The skin begins to break apart. Without the restoration of blood flow, the outer layers of the skin begin to die. This damage is easily noticeable and may appear as shallow craters in the skin. At Stage 2, the pressure ulcer is painful and very inflamed. There may be blister-like wounds and pus. Stage 3: The tissue decay progresses past the first two layers of the skin. The sore is now an open wound. The wound may have a foul odor and be consistently oozing. The skin and tissue may have discoloration. Stage 4: The wound has reached all the way through the skin to muscle, bone, or tendons underneath. At this stage, the wound is necrotic and has a foul odor and an abundance of pus or drainage. The wound is deep enough for sepsis to be a significant risk. Without immediate medical care, these pressure ulcers can be life- threatening.

Treatment

Venous Ulcers

A venous skin ulcer is a shallow wound that occurs when the leg veins don't return blood back toward the heart the way they should. This is called venous insufficiency. These ulcers usually form on the sides of the lower leg, above the ankle and below the calf. Venous skin ulcers are slow to heal and often come back if you don't take steps to prevent them. A venous skin ulcer is also called a stasis leg ulcer.

Cause

Venous ulcers most often form around the ankles. Venous ulcers typically occur because of damage to the valves inside the leg veins. These valves control the blood pressure inside the veins. They allow it to drop when you walk. If the blood pressure inside your leg veins doesn't fall as you're walking, the condition is called sustained venous hypertension. That increase in blood pressure causes ulcers to form on your ankles. Venous skin ulcers are caused by poor blood circulation from the legs, such as from venous insufficiency. Your veins have one-way valves that keep blood flowing toward the heart. In venous insufficiency, the valves are damaged, and blood backs up and pools in the vein. Fluid may leak out of the vein and into the surrounding tissue. This can lead to a breakdown of the tissue and an ulcer. Veins that become blocked also may cause fluid to pool, leading to these ulcers. Some things can increase your risk of venous skin ulcers. These include: Deep vein thrombosis, in which a blood clot (thrombus) forms in the deep veins of

the legs. Obesity. Smoking. Lack of physical activity. Work that requires many hours of standing.

Cause

Venous ulcers may also be caused by other problems with your leg veins. These include: Varicose veins. These are large, bulging leg veins. They occur when valves in the leg veins don’t work well, allowing blood to collect (pool) in the lower leg. Chronic venous insufficiency. Like varicose veins, this condition occurs when your leg veins can’t pump blood back up to your heart. Blood then pools in your lower legs, causing your legs to swell. Since the blood can't flow well in your legs, the swelling may be extreme. This extreme swelling can put so much pressure on your skin that venous ulcers form.

Treatment

The four phases of wound healing

The complicated mechanism of wound healing occurs in four phases: hemostasis, inflammation, proliferation, and remodeling. Hemostasis, which occurs just after injury, utilizes clotting factors which prevent further blood loss from the wound site as well as the structural foundation for the future formation of granulation tissue. The subsequent inflammation phase, involving phagocytic cells that release reactive oxygen species, may last for up to seven days in acute wounds and longer in chronic wounds. As inflammatory cells undergo apoptosis, wound healing progresses to the proliferation phase, which is characterized by the formation of granulation tissue, angiogenesis (blood vessel formation), wound contraction, and the process of epithelialization. The final remodeling phase, characterized by the formation of scar tissue, may occur over a period of months or years, depending on the initial severity of the wound, location, and treatment methods.

Infected wound healing stages

Granulation tissue, composed of endothelial cells, capillaries, keratinocytes, and fibroblasts, is also an important component of wound healing. This connective tissue can provide important indicators of wound healing progress; pink granulation tissue is considered to be healthy and a sign that healing is progressing normally, while dark red tissue may be a sign of infection. Bacterial Overgrowth of granulation tissue, characterized by a white or yellow film, is seen occasionally in infected wounds and must be removed before healing can continue. Wound healing is a multifactorial process involving blood cell coagulation, inflammatory cell response, and granulation tissue formation. Several factors and conditions may contribute to the occurrence or persistence of a chronic wound, such as weakened immune function, comorbidities, venous insufficiency, and lack of proper circulatory function. The immune system plays an integral role in wound healing by mobilizing stem cells, promoting cell differentiation, and stimulating growth factors which ultimately result in angiogenesis, or the formation of new blood vessels. When immune activities are disrupted through medications or comorbidities, the process of wound healing may become stalled, leading to persistent or chronic wounds.

Chronic wounds do not follow the standard progression of wound healing seen in acute wounds, and instead tend to arrest temporarily in one of the wound healing phases (most commonly the inflammation phase). The healing process of an infected wound may also be prolonged compared to that of a non-infected wound. In infected wounds, pathogenic organisms enter the wound tissue and disrupt normal skin flora, leading to increased inflammation and damage of sensitive new tissue growth. While some infected wounds may resolve without intervention, in order to accelerate the wound healing process and ensure further complications (such as cellulitis, osteomyelitis, or septicemia) do not occur, infected tissues should be treated as soon as possible. Treatment of an infected wound differs in some ways from that of a non-infected wound, as it involves eliminating the infection with oral or topical antibiotics, draining or debriding the wound to remove dead tissue, and applying antimicrobial dressings.

Factors that Affect Wound Healing

There are several factors that may impair the wound healing process, including: the pre-existing integrity of the wounded skin due to age or medical treatments, comorbidities, medications, infection, hydration state, nutritional status, lifestyle habits, and pre- and post-operative care if surgery has occurred. Relevant comorbidities, medications, and lifestyle factors include: Diabetes: A common complication associated with diabetes is peripheral neuropathy leading to foot ulceration. An additional complication is peripheral ischemia secondary to peripheral artery disease. Both complications affect the proliferative phase of healing and lead to the overall slowing of wound healing. Obesity: Obesity is associated with an increased risk of ischemia and inadequate tissue oxygenation, which may lead to slowed wound healing or necrosis. Necrosis: Unplanned tissue death is another factor that may impede wound healing, requiring debridement to remove the affected tissue surgically before healing can proceed. Poor nutrition: Malnutrition (seen frequently in elderly patients), specifically inadequate protein intake, can lead to decreased blood vessel formation, collagen production, and fibroblast proliferation, which ultimately slows the wound healing process. NSAIDs (non-steroidal anti-inflammatory drugs): The mechanism of pain reduction by NSAIDs occurs through the inhibition of PGE2, an inflammation mediator. NSAIDs are known to slow wound healing through the halting of angiogenesis. NSAIDs also increase scar formation, particularly if used during the proliferative phase. Steroids: The anti-inflammatory and immunosuppressive effects of steroids can hinder wound healing by decreasing fibroblast proliferation and collagen production.

Radiation therapy: Ionizing radiation beams can damage epithelial cells as they pass through to targeted tissues, causing skin tissue breakdown and slowed healing of existing and new wounds. Chemotherapy: Chemotherapeutic agents affect wound healing by delaying the inflammatory phase of healing and decreasing collagen production. Smoking: Cigarette smoking, specifically the use of nicotine, affects blood flow by causing vasoconstriction. Nicotine also decreases the body’s immune response, which could lead to an increased likelihood of wound infection. Alcohol: Alcohol intake is often associated with poor nutritional habits, which may result in decreased immune function. In addition, alcohol may impair wound healing by decreasing angiogenesis and collagen formation, leading to weaker scar tissue formation and an overall slower healing process. Wound healing may also be positively impacted by the addition of certain supplements such as zinc and vitamin C.

How chronic and acute wounds heal

While acute wounds typically follow the normal healing process of hemostasis, inflammation, proliferative tissue regrowth, and tissue strengthening through remodeling, chronic wounds tend to progress through the healing process at a slower pace. Healing of a chronic wound may arrest for several weeks in one of the four phases–most commonly the inflammatory phase. There are several known factors that affect the mechanism of chronic wound healing. These factors include: the presence of inflammatory cytokines or growth factors, infection at the wound site, formation of a biofilm over the surface of the wound, hypoxia (often associated with cardiovascular, pulmonary, and vascular diseases), and a nutrient-poor diet. Research suggests various methods to accelerate the wound healing process, such as ensuring adequate sleep to minimize the effect of pro-inflammatory cytokines, adding nutrient-rich foods to a patient’s diet (foods that contain vitamin A and zinc are particularly helpful), avoiding cigarette smoking, and regularly cleaning and dressing the wound. Once a wound has healed and begun the scarring process, the American Academy of Dermatology recommends applying petroleum jelly to the wound site to minimize dehydration of the scar and surrounding tissue, as well as applying sunscreen to the site daily to reduce hyperpigmentation associated with scar tissue.

Amniotic tissue products are powerful options for physicians to keep in their wound care arsenal. These products offer the potential for accelerating wound healing, reducing the incidence of infection and preventing reulceration. Yet, despite the published advantages of using amniotic membrane in treating DFUs, it is not appropriate to apply them to every wound. Advanced tissue products are not a replacement for the traditional mainstays of wound care such as aggressive sharp debridement, adequate offloading and the application of sterile dressings. Amniotic membrane, like most biologic tissue products, requires significant processing and therefore carries a high price tag. On average, amniotic tissue products cost $500 to $1,000 per application. This may seem astronomically high at first glance but the treatment and complications of DFUs carry an approximate cost of $28,000 per patient per year.17 Therefore, when clinicians use amniotic membrane products appropriately, the modalities may result in sizeable cost savings for our health care system. Furthermore, in the setting of treating patients who are at a high risk for amputation, financial considerations must take a back seat to limb preservation efforts. Further studies are needed to determine whether improved healing rates achieved with these products correlate with a reduced rate of amputation and mortality.

Treatment for Wound Care

Homologous Use Definition: Repair, reconstruction, replacement, or supplementation of a tissue with an HCT/P that performs the same basic function in the recipient as in the donor

Amniotic Membrane & Fluid

Pluripotent Cells- can differentiate into any mature cell type from the 3 germ layers:

Endoderm (lungs, GI tract, interior stomach lining) Mesoderm (bones, muscles, blood, and urogenital tract) Ectoderm (epidermal tissues and nervous system)

Multipotent Cells- can differentiate into a limited number of mature cell types (e.g. hematopoietic blood-forming cells)

(Delo et al., Amniotic Fluid and Placental Stem Cells, Methods In Enzymology, Vol. 419 (2006)

Amniotic Membrane

Amniotic membrane has a number of characteristics that make it especially suited to wound healing. The amniotic membrane: Sterile allograft derived from amniotic membrane Fully resorbed in body during healing process Simple, suture less technique for placement at site contains a significant number of cytokines and essential growth factors reduces pain when applied to a wound increases and enhances the wound healing process has antibacterial properties is non-immunogenic (will not be seen as foreign material) provides a biological barrier provides a matrix for migration and proliferation of cells reduces inflammation reduces scar tissue formation

Wound Healing Properties of Amniotic Membrane

Amniotic Membrane - Key Components

Sterile allograft derived from amniotic membrane Fully resorbed in body during healing process Simple, suturel ess technique for placement at site Provides a natural substrate for facilitating reparative cell migration, adhesion, differentiation and maintenance Non-immunogenic Reduces inflammation Reduces pain at the site of application May reduce scar tissue adhesions Creates a protective wound barrier Contains useful growth factors and cytokines

Dehydration of Amniotic Membrane Tissue

In the past, amniotic tissue was sterilized and stored at 4°C. Amniotic tissue could only be used for up six weeks, at which point it was no longer useful. Now, this material can be cleansed, dehydrated and sterilized, which means that the shelf life of amniotic membrane has been greatly increased. Indications for Use What types of wounds can amniotic membrane be used on? Traditionally, amniotic membrane has been used on burns; nowadays, however, amniotic membrane can be used on a wide variety of wounds. It is important to note that amniotic membrane should be used only after conservative treatment has failed. In other words, amniotic membrane may be used for wounds that are chronic and non-healing.

Application of Amniotic Membrane

Prior to using amniotic membrane, a thorough initial assessment of the wound is necessary, as is a medical history. Gather information on the history of the wound, including duration, what treatments have been tried and patient comorbidity. Document wound appearance, size, depth, presence of necrotic tissue and note whether bone or other structures are visible. Assess circulatory status, nutrition and other barriers to healing. These are the same steps that you would follow prior to implementing any wound treatment and are not specific to amniotic membrane treatment. Next, prepare the wound bed by performing any necessary debridement. The wound bed should be clear of any necrotic tissue and should not have any signs of infection. Amniotic membrane is supplied in a sterile container, and sterile scissors may be used to cut a piece of the amniotic membrane to fit the wound. The material may be applied wet or dry. Note that the stromal collagen layer must be facing the wound - read the manufacturer's recommendations to determine how to apply the material. You can use steri-strips to hold the graft in place. There is no need to suture the material in place. A secondary dressing which promotes moist wound healing should be chosen as a secondary dressing. The graft should not be disturbed for at least one to two weeks. After one to two weeks the amniotic membrane allograft will be incorporated into the wound. You should begin to see improvement in the wound in terms of size and depth within 2 to 3 weeks, or even sooner. You can apply a second graft once slowing of wound healing has occurred, as typically observed by wound measurements over time (Podiatry Today, 2015). The use of amniotic membrane in the management of chronic wounds is an exciting new development which provides another option for wounds that fail to heal using traditional wound therapies and dressings.

Amniotic Fluid - Key Components

The primary function of amniotic tissue for implant is to provide a structural tissue matrix to fill, cover and protect a wound or defect, the presence of viable cells may provide ancillary clinical benefits by enhancing the body’s natural regenerative process Amniotic tissue matrix includes the collagens, proteins, lipids, carbohydrates, hyaluronic acid, growth factors and other chemical compounds that are needed for tissue growth and healing The primary function of amniotic tissue for implant is to provide a structural tissue matrix to fill, cover and protect a wound or defect the presence of viable cells may provide ancillary clinical benefits by enhancing the body’s natural regenerative process Anti-inflammatory Cryopreserved, multipotent tissue matrix Amniotic tissue is an abundant source of collagen that provides an extracellular matrix to act as a natural scaffold for cellular attachment Amniotic membrane and fluid is immune privileged, class II antigens are NOT expressed

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Amniotic Fluid

Chronic Cellular tissues can migrate, proliferate and differentiate. This process can provide a benefit by helping recruit those cells that are needed in order for healing to occur at the wound site.

Amnion Fluid Preparation

Amniotic fluid is maintained at -65°C or colder until it is ready for use

DO NOT REMOVE AMNIOTIC FLUID FROM THE DRY ICE OR FREEZER UNTIL YOU ARE READY TO PREPARE IT FOR USE

Remove product from freezer or container. Verify expiration date and serial number Allow amnion fluid to thaw for about 5 minutes by hand or 8-10 minutes by leaving it on a table

THE EXTERIOR POUCH AND THE OUTSIDE OF THE VIAL CONTAINING THE AMNIOTIC FLUID ARE NOT STERILE DO NOT DROP ON A STERILE FIELD

Preparation Continued

Amniotic Peel open the outer package and remove the inner pouch Peel open the inner pouch utilizing aseptic technique Once thawed, unscrew the top of the vial and either draw up the product with a sterile needle (18g or larger) and syringe

Preparation Continued

Dilute Cryopreserved allograft with Any of the following: Normal saline 1% preservative free plain lidocaine Platelet Rich Plasma Platelet Poor Plasma Mix using 18-gauge needle in 3 / 5/ 10 ml syringe Use Nothing smaller than 23-gauge needle for injection Dilute in a 1:1 ratio is typical You can dilute in higher ratios if need more volume.

Preparation Continued

If there is to be a delay in the use of amniotic fluid, it is recommended that it be immediately prepared with the recommended dose of lidocaine which will help prevent the cells from dying. However, the maximum delay should not exceed 15 minutes Do not dilute amniotic fluid with any epinephrine or glycerol products

Amnion is Anti-Inflammatory

Amniotic membrane cells express various anti-angiogenic and anti-inflammatory proteins (IL- 4 and IL-10, IL-1 receptor antagonist 2, and tissue inhibitors of metalloproteinase i.e. TIMPs -1 thru 4 3) Amniotic membrane contains anti-inflammatory factors that may enhance wound healing 1

1. John T. et al Human amniotic membrane transplantation: Past, present, and future. Ophthalmology Clinic of North Am. 2003;16:43-65 2. Toda A. et al The Potential of Amniotic Membrane/Amnion-Derived Cells for Regeneration of Various Tissues. J. Pharmacol Sci 2007;105:215-228 3. Hoa Y, Ma DKH and Hwang DG. Identification of ant-iangiogenic and anti- inflammatory proteins in human amniotic membrane. Cornea 2000;19: 348-352

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Amnion is Non-Immunogenic

Acute immune rejection does not occur after the transplantation of human amniotic epithelial cells 1 No detectable immune response to amniotic membrane 2 Dry amnion does not contain viable cells The tissue does not produce HLA class I antigens (HLA-A, B, C , G, and E) or β -2 macroglobulin Contains immunomodulatory cytokines IL- 4 & 10, and TGF- β 1 & 2 The epithelial layer contributes significant immunosuppressive properties with dehydrated amniotic membrane products

1. Immunosuppressive Properties of Dried Human Amniotic Membrane 2007 ARVO abstract #B737 2. Akle, C.A et al. Immunogenicity of Human Amniotic Epithelial Cells after Transplantation into Volunteers. The Lancet. 1981;2(8254);1003-1005.

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Scar Reduction

Amniotic membrane inhibits fibrosis

Inhibits expression of TGF- b receptors in fibroblasts resulting in less fibrosis1 Serves as an anatomical barrier that can block fibrosis2

1. Immunosuppressive Choi TH, Tseng SCG. In vivo and in vitro of epithelial cell-induced myofibroblast differentiation of keratinocytes and limbal fibroblasts by amniotic membrane matrix. Cornea 2001;20:197-204 2. John T. et al Human amniotic membrane transplantation: Past, present, and future. Ophthalmol Clin North Amer 2003;16:43-65

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Amnion is Non-Immunogenic

Collagen Types IV, V and VII Several growth factors

Epidermal Growth Factor (EGF) Transforming Growth Factors-beta (TGF-b 1&2) Fibroblast Growth Factors (FGFs) Platelet Derived Growth Factors (PDGF A&B)

See: Hopkinson A. et al Proteomic Analysis of Amniotic Membrane Prepared for Human Transplantation: Characterization of Proteins and Clinical Implications. Journ Proteome Res. 2006; 5:2226-2235. Confidential

Amnion

Chorion

The Wound Healing Process

Step 1: Inflammatory Phase (Immediate to 2-5 days) Hemostasis Inflammation Step 2: Proliferative Phase (2 days to 3 weeks) Granulation Fibroblasts lay bed of collagen Contraction Wound edges pull together to reduce defect size Epithelialization Step 3: Remodeling Phase ( 3 weeks to 2 years) New collagen forms which increases tensile strength to wound

Amnion contains anti- inflammatory cytokines, TIMPS

Amnion provides collagen substrate & growth factors that support wound healing

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Preparation

Remove Product is to be stored at ambient temperature Remove package from the box Peel open the outer envelope utilizing aseptic technique (the inner pouch is sterile) Open the inner pouch and pass to the sterile field utilizing aseptic technique Remove one side of the mesh to expose the membrane After placing the membrane on the desired site, remove the remaining mesh DO NOT LEAVE THE MESH IN THE PATIENT Confidential

Case Studies

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Patient 1 Comparative

Day 189 – Closed Margin

Day 0 – 5cm x 4cm

Diabetic Ulcer

Inhibits Initial Procedure Date: 17 Mar 2011 History:

49 yr, african american, male, diabetic; renal failure; dialysis x3/week; severe lymphodema; awaiting kidney transplant; suffers from two (2) year non-healing right foot ulcer preventing application for transplant; 5th metatarsal removed; four (4) doctors previously recommended amputation – patient denied recommendation each time. Initial Treatment: 1. Ulcer debridement 2. Two (2) 1.25 ml amnion diluted with plain lidocaine 1:1 and injected sc/sq around ulcer margin at eight unique points into wound interior.

Patient 1 Pre-debridement

Day 189 – Closed Margin

4.9cm margin

Deep sinus (not visible) poses major threat to foot

Patient 1 - Day 5 Follow Up - Inflammation

Day 5 - Inflammation reduction

Day 0

Patient 1 - Day 21

Day 21 Inflammation reduced & improved color/vascularity

Day 189 – Closed Margin

Deep Visible ankle structure

Patient 1 - Day 21 Comparative

Day 21 – 1.7 cm margin

Day 0 – 2.5 cm margin

Patient 1 - Day 57 Comparative

2x2cm acellular skin matrix

Capillary bed ready for graft

Adaptec applied over skin graft

Patient 1 - Day 63 Margins Closed

Patient 1 - Day 21 Comparative

Day 21 – 1.7 cm margin

Day 0 – 2.5 cm margin

Patient 1 Margins Closed

Back on transplant list Back at work

Received Kidney!!!

Patient 2 - Pressure Ulcer History: 87 yr old, nursing home, immuno-compromised, renal insufficiency, diabetic, non- ambulatory patient suffers from a nine (9) month non-healing left heel pressure ulcer. Protocol: 1. Ulcer debridement 2. One( 1) 1.00 ml amnion diluted with plain lidocaine, 1:1 3. Injected sc/sq around ulcer margin at 5 unique points.

4. 2x2cm amniotic membrane tacked in 5. Telfa™, non-porous cover, Coban™ wrap

Patient 2 – Day 0 Allograft Implant

Day 0 - Initial view amnion fluid 1.0ml by injection

Patient 2 – Day 0 AGGRESSIVE DEBRIDEMENT

2.4 cm x 2.4 cm margins

Single 2x2 cm amniotic membrane patch

Patient 2 – Day 13 Comparative

Day 13: depth=1.5 mm Margin= 0.8 cm

Day 0: depth=3 mm Margin=2.4 cm

Patient 2 – Day 55 Follow-up

Significant epithelialization and granulation Note: Non-compliant patient missed two scheduled appointments (30 days) due to failing health, including diminished renal function

Patient 2 – Day 70 Margins closed

Patient 2 – Day 83 Margins closed

Wound 1: left foot 2nd metatarsal ulcer Wound 2: right foot 1st metatarsal draining ulcer Wound 3: single right foot bunion ulcer

Patient 3 History: 69 yr, female, diabetic, immunosuppressive agents for 01/2010 kidney transplant. Exam indicates peripheral neuropathy. Wound #1 is a left foot 2nd metatarsal draining ulcer; wound #2 is a right foot 1st metatarsal draining lesion wound #3 is a right foot bunion lesion.

Protocol: 1. Minor ulcer debridement

2. Two (2) 1.00 ml amnion fluid diluted with saline 1:1 and injected sc/sq with equal distribution around all three (3) ulcer margins and into each wound bed. 3. Telfa™ and non-porous wound covering applied

Patient 3 - Day 0 Wound 1 (left 2nd met.)

1.8 cm margin

1.5 cm margin

Patient 3 - Day 8 Wound 1 – Follow-up

Patient 3 - Day 43 Wound Closed

Patient 3 – Day 0 Wound 2 (rt. ft 1st met.)

0.8 cm margin

1.1 cm margin

Patient 3 - Day 36 Wound 2 - Closed

Patient 3 - Day 0 Wound 3 (rt. ft bunion lesion)

1.3 cm margins

1.4 cm margins

Patient 3 - Day 7 Wound 3 (rt. ft bunion lesion) Day 7: X-ray indicates possible osteomyelitis beneath bunion Day 0: Patient injected lesion

- Patient is scheduled for osteotomy in 2 weeks. Day 14 and Day 21 : X-rays indicate that possible osteomyelitis is resolving - Patient consents to continued monitoring by X- ray. (Note: Patient is a high- risk surgical candidate. Both doctor and patient are making best efforts to avoid surgical intervention)

Patient 3 - Day 43 Wound 3 - Follow-up Note granular tissue and “buds”

Patient 3 - Day 57 Wound 3 – Margins Closed

Patient 3 – Day 57 Wound 3

Day 7 Day 57: Bone fill with slight loss of outer cortex

Patient 4 History: Wound #1 is a trauma-induced

Left, distal, lateral lower leg pressure ulcer

(28 Aug 2010) pressure ulcer located on left, distal, lateral lower leg caused by trauma/impact nearly 7 months prior. Patient is a 59 yr African-American; smokes 2 ppd, 71”, 265#. Good pulse; circulation good by imaging. Pain level 9 is controlled by Percocet® (5mg oxycodone/ 325mg acetominophen), 6-8 q.24/hr. Margins are increasing. Initial Protocol: 1. Left, distal, lateral lower leg pressure ulcer : Minor ulcer debridement 2. Two (2) 1.00 ml amnion fluid diluted with saline 1:1 and injected sc/sq around ulcer margin at 12-16 unique points and into wound bed. 3. Four (4) 2x6cm amniotic membrane allograft 4. Telfa™ and dry wound covering 5. Coban™ wrap

Patient 4 - Day 0

Patient 4 - Day 161 Comparative

Day 0 (6cm x 4cm) (12cm x 10cm) Vascular bed ready for graft

Day -161

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