Dressing

Abstract

Described herein are surgical or wound dressings comprising a sheet of gellan gum and an antifibrotic agent.

Methods for producing dressings comprising gellan gums and biologically active agents are also described.

Claims

1.-36. (canceled)

37. A method of producing a surgical or wound dressing comprising a gellan gum and an antifibrotic agent, the method comprising: (a) forming a layer of the gellan gum onto a mold; (b) depositing a layer of the antifibrotic agent onto the layer of gellan gum; (c) forming a further layer of heated gellan gum onto the layer of antifibrotic agent; (d) optionally repeating steps (b) and (c) one or more times; and (e) optionally depositing a layer of antifibrotic agent onto the final layer of gellan gum.

38. The method of claim 37 wherein the heated gellan gum is about 0.05 w/w to about 5% w/w gellan gum in an aqueous liquid.

39. The method of claim 37 wherein the antifibrotic agent comprises decorin.

40. A dressing produced by the method of claim 37.

41. The dressing of claim 40 wherein the dressing is dried after molding.

42. The dressing of claim 41 wherein, before use, the surgical dressing is rehydrated.

43. The dressing of claim 40 additionally comprising on at least a portion of a surface of the dressing, a region comprising one or more proscarring compounds.

44. The dressing of claim 40 further comprising a chemically modified surface for the attachment of one or more cells, or peptides, or a combination thereof.

45. A method of treating an eye, wound, burn, or surgical incision, the method comprising applying the dressing of claim 40 to the eye, wound, burn, or surgical incision.

46. The method of claim 45 wherein the surgical incision is an incision in the brain, abdomen, the socket of a tooth, a tendon or ligament, the skin, or a wound from the removal of one or more tumors.

47. A bandage or sticking plaster comprising a layer of the dressing of claim 4.

48. A method of making a surgical or wound dressing, comprising heating together: (a) about 0.05 to about 5% w/w gellan gum and an aqueous liquid optionally with 0 to about 5% w/w of a further polymer, to form a liquid gellan gum; (b) cooling the liquid gellan gum; (c) adding an antifibrotic agent to the cooled liquid gellan gum; (d) casting the gellan gum to form a sheet of gellan gum having a thickness of about 0.5 to about 5 mm thickness; and (e) optionally drying the sheet of gellan gum.

49. The method of claim 48 comprising mixing the gellan gum and aqueous liquid at about 80 C. or greater to form liquid gellan gum; cooling the liquid gum to about 37 C. to about 45 C.; mixing an antifibrotic agent in the cooled liquid gellan gum at about 37 C. to about 45 C.; and cooling and casting the liquid gellan gum containing the antifibrotic agent.

50. The method of claim 48 wherein the dried sheet of gellan gum is rehydrated with an aqueous liquid, wherein optionally the aqueous liquid comprises one or more biologically active additives.

51. The method of claim 50 wherein the rehydrated dressing is compressed to exclude a portion of liquid from the dressing; the excluded liquid is removed; liquid comprising one or more biologically active additives is placed in contact with the dressing; and the dressing is allowed to expand to draw the liquid containing the biologically active additive into the wound dressing.

52. The method of claim 50 wherein at least one of the biologically active additives is selected from the group consisting of antifibrotic agents, pro-VEGF agents, anti-VEGF agents, antibacterial agents, and proteoglycans.

53. The method of claim 48 further comprising contacting the cast gellan gum with a cross-linking agent.

54. A method of analyzing a condition of a wound or burn, the method comprising contacting the wound or burn with the dressing of claim 40, allowing fluid from the wound or burn to pass into or through the dressing, and analyzing the fluid in the dressing or which has passed through the dressing.

55. A dressing comprising at least one layer of an antifibrotic agent and at least two layers of a gellan gum, where at least one layer of the antifibrotic agent is positioned between two such layers of the gellan gum.

56. The dressing of claim 56 wherein the dressing is capable of or adapted for use in treating an eye, wound, burn, or surgical incision.

Description

[0073] The invention will now be described by way of example only with reference to the following figures:

[0074] FIG. 1 shows a hydrogel dressing sheet manufactured according to the invention.

[0075] FIG. 2 shows the effect of a variety of calcium chloride concentrations on the G of gellan membranes.

[0076] FIG. 3 compares the release of decorin, decorin from gellan membranes and amniotic membranes. FIG. 3a shows the difference between gellan only and gellan cross-linked membranes. FIG. 3b shows the release from amniotic membrane.

[0077] FIG. 4 shows an in vitro assessment of gellan membranes and shows the collagen deposition from MFs in the presence of gellan membranes, and, which have been assayed using sirius red.

[0078] FIG. 5 shows that decorin significantly reduces the amount of collagen deposited over a 12 day period when compared to non-treated samples.

[0079] FIG. 6 shows a schematic diagram of the surface modification of gelatin to allow the attachment of MF cells.

[0080] FIG. 7 shows the effect of adding PVA (polyvinyl alcohol) to gellan.

MATERIAL AND METHODS

[0081] Industrial grade gellan was obtained from Kelco Limited (Surrey, United Kingdom) under the trade name Gelzan. Calcium chloride, and phosphate buffered saline (Dulbeccos A) were obtained from Sigma Aldridge Company, Dorset, United Kingdom. PETRI dishes, filter paper range (QL100-240 mm) and disposable 10 mm plastic syringes were obtained from Fisher Scientific, Loughborough, United Kingdom. The incubator used by an INCU-LINE, obtained from VWR, Sussex, United Kingdom.

[0082] 2% w/w gellan hydrocolloid solution was prepared by weighing 2 grams of gellan powder using a calibrated balance. 98 ml of distilled water was measured and transferred to a glass bottle. Water was placed on a hot plate stirrer at 100 C. and 200 rpm. Gellan powder was added to the stirring water and left for four hours.

[0083] A 1% w/w calcium chloride solution was prepared by dissolving 1 gram of calcium chloride in 99 ml of distilled water.

[0084] The gellan hydrocolloid was stirred on a hot plate at 90 C. until the gellan became a liquid. This was then reduced to 40 C. 2.5 ml of gellan was syringed into a 55 mm petri dish using a plastic syringe. This was allowed to set for 15 minutes. To dehydrate the membrane sheet, the gel was placed in a 37 C. incubator for 6-8 hours.

[0085] To prepare a cross-linked sheet, the gelled gellan hydrocolloid was stirred on a hot plate at 90 C. at 200 rpm, until the gellan became liquid. This was reduced to 40 C. A filter paper was soaked in the calcium chloride solution and placed on the bottom of a petri dish. 2.5 ml of gellan was placed onto the filter paper using a plastic syringe. The gel was allowed to set for 5 minutes. A further filter paper soaked in the calcium chloride solution was placed on top of the set gel and left at room temperature for 15 minutes. The filter papers were then carefully removed and the membrane sheet was transferred into an incubator at 37 C. for 6-8 hours.

[0086] In order to rehydrate the sheets, the sheets were soaked in PBS for 2-3 minutes.

[0087] FIG. 1 shows an example of the dehydrated sheet.

[0088] The Effect of Various Calcium Chloride Concentrations on G Gellan Membranes

[0089] Gels were prepared with a range of different concentrations of calcium chloride. 276 mM calcium chloride, the gel was stiff, not easily drapable or easy to place around a wound. Gels prepared with 15 mM were shown to be more drapable around a spherical object, such as a marble, but were susceptible to tearing at maximum strain. 10 mM calcium chloride were more fragile and would tear more easily when lifted out of the mould. Therefore 15 mM calcium chloride was the preferred concentration to use. The effects on G are shown in FIG. 2.

[0090] FIG. 3 shows a comparison of the release of decorin from gellan membranes and amniotic membranes. Gellan-decorin membranes or amniotic membranes were placed in a release medium (Phosphate buffer saline, PBS) and placed in a shaker at 80 rpm at 37 C. Equal amounts of the release medium were taken at specific time intervals and equal amounts of fresh release medium were replaced to maintain a constant volume. The samples were analysed using a human decorin ELISA kit according to manufacturer's instructions

[0091] FIG. 4MFs were cultured in 6-well plates and left to attach overnight. After overnight attachment TGF-1 was added to the cells to produces excess collagen. Decorin containing membranes were placed on top of the cultures and collagen production was observed using the Sirius Red assay over time.

[0092] FIG. 5: For quantifying the amount of Sirius red dye bound to collagen in the cultures the Sirius red was dissolved with 0.1N sodium hydroxide and left on a shaker for 30 min at room temperature. The dissolved dye was measured colourimetrically using a plate reader at 550 nm

[0093] FIG. 6: Gellan membranes were produced and cross-linked with 200 mM calcium chloride, and immersed in 10 mM sodium periodate solution for 2 h. The membranes were washed with PBS and further immersed in 10% (w/v) gelatin and 0.5% (v/v) NaBH.sub.4. Furthermore the membranes were soaked overnight in EDC at 4 C.

[0094] To determine surface modification, MF cells were left to attach on the surface on membranes for 24 h and cell attachment was assessed using Calcein AM and Propidium Iodide staining.

[0095] FIG. 7: Gellan membranes were produced with various amounts of PVA at 10:0, 50:50 and 20:80 (wt %) of gellan:polymer ratios. Tensile testing was carried out using a universal Instron machine to determine the physical properties of the gellan/PVA membranes

[0096] Method of Spray Forming a Gel Dressing

[0097] 2% w/w of gellan was dissolved in deionised water. A layer of the gellan gum at 60 C. was sprayed onto a mould. After spraying a layer of dextran blue, which was used as a substitute for decorin because it has similar properties and is visible to the naked eye, was sprayed onto the layer of gellan after a 5 second interval to allow the gellan to cool and gel. A 5 second interval was allowed prior to adding another layer of gellan gum and the process repeated 5 times. The whole process took approximately 40 seconds.

[0098] It was found that it was possible to form a structure with alternating layers of gellan and dextran blue, which was membrane-like. The advantage of this system is that it allows the material to be rapidly produced. Rastering the spray backwards and forwards across the surface allows a substantially even distribution of the material. Dextran blue was substantially evenly distributed within the gel dressing.

[0099] The advantage of this system is that it improves the ability to produce the dressing. It also lends itself to automation in that the spraying of the materials can be readily automated.