![Influence of hyaluronan (HA) preparations on ionizing-
radiation-treated collagen-based tissues
Huber A.T. BS, Meyer A.E. PhD, Zhao R.G. PhD, Baier R.E. PhD
Biomaterials Graduate Program, SUNY at Buffalo, Buffalo, NY, USA
Abstract #__
ABSTRACT
RESULTS
CONCLUSIONS
INTRODUCTION
MATERIALS AND METHODS
REFERENCES
ACKNOWLEDGMENTS
Objectives:
It has been proposed that HA solution (.5% HA in distilled water, DW), due to its natural lubrication properties [1], could remedy
xerostomic patients. Investigation was conducted using Tensile, Friction, and Hydroxyproline (HYP) Assay Studies. The experimental
aim was to observe effects of 70 Gray 稗-Radiation (standard dose for head/neck cancer patients) and HA-soaking.
Materials & Methods:
Tensile: 19 non-irradiated specimens of fresh human oral mucosa (OM) and bovine pericardium (PC) and 35 irradiated specimens
were soaked in DW or HA. Tensile testing until failure was conducted.
Static Friction: 13 PC/OM samples were tested before and after 粒-irradiation, 7 DW-soaked, 6 HA-soaked. Tissues fastened to friction
device, limiting slippage.
HYP Assay: OM/PC samples, soaked in HA/DW, Irradiated and Non-irradiated, were tested. Developing a standard curve involved:
Digestion of samples in HCl at 105 属C and adding a color reagent. Sample %Transmittance was compared to standard curve.
Results:
Tensile: Non-Irradiated PC/OM tissue had higher average strain (0.825 +/- 0.311 vs 0.485 +/- 0.186) and lower ultimate strength (9.858
+/- 1.528 MPa vs 13.597 +/- 4.480 MPa) than Irradiated PC/OM. PC had a slightly higher average strain of 0.726 +/- 0.124 and 13x-higher
average ultimate strength of 21.818 +/- 6.398 MPa than OM.
Static Friction: OM HA had about 遜 the 亮 (friction coefficient) as OM DW pre-irradiation. After irradiation, OM HA and DW 亮s merged
to 0.45. PC HA had about 遜 the 亮 as PC DW before/after irradiation.
HYP Assay: HCl-digestion-based HYP Assay is equally capable of hydrolyzing to their AA contents BOTH natural and irradiation-
crosslinked tissues[2]. Final test will find if HCl-undigested samples will determine if irradiation had released any HYP due to collagen
chain scission. Preliminary findings suggest that tissue degradation of OM is accelerated when irradiated and therefore releases
more HYP when pulped, hinting at collagen chain scission being more abundant than chain crosslinking. HAs possible protection
against this chain-scission is still being evaluated.
Conclusion:
HA solution relaxes the strain of OM. HA shields OM/PC during irradiation, which otherwise causes tissue embrittlement. HA
significantly reduces friction for irradiated/non-irradiated PC/OM. Irradiation increases degradation of tissue, while increasing its
elastic modulus and increasing its 亮.
It has been observed that HA provides superior and reversible physical protection to collagen-rich tissues as monitored in
friction testing. Considering the possible benefits[3] to irradiated head-and-neck cancer patients who have lost all natural
salivary lubrication, fresh human oral mucosa was collected and tested against a crosslinked standard pericardium reference
material for its ability to take up HA reversibly and preserve desirable tissue properties after exposure to clinically-used doses of
70 Gray (of Cs-137) 稗-irradiation.
Adding HA relieved the tensile strain of both normal and irradiated samples, and provided some modulation of the radiation-
induced crosslinking and embrittling (increase in Elastic Modulus) of OM. Weight measurements illustrated that HA was taken up
into native and irradiated tissues, and was completely reversible by water exposure, so the effect was more than superficial
lubrication (which lasts only short times). Therefore, the predominant mechanisms of protection of HA formulations are friction
reduction in both irradiated and non-irradiated OM and strain relief, which now remain to be correlated with subjective pain relief
and improved oral function. This mechanical disparity correlates with OM having 1/3 the collagen amount as PC.
[1] Singh A, Corvelli M, Unterman SA, Wepasnick KA, McDonnell P, Elisseeff JH. Enhanced lubrication on tissue and biomaterial surfaces through peptide-
mediated binding of hyaluronic acid. Nat Mater. 2014;13:988-95.
[2] Bailey AJ, Rhodes DN, Cater CW. Irradiation-Induced Crosslinking of Collagen. Radiation Research. 1964;22:606-21.
[3] Kawano T, Miura H, Mawatari T, Moro-Oka T, Nakanishi Y, Higaki H, et al. Mechanical effects of the intraarticular administration of high molecular
weight hyaluronic acid plus phospholipid on synovial joint lubrication and prevention of articular cartilage degeneration in experimental osteoarthritis.
Arthritis & Rheumatism. 2003;48:1923-9.
I would like to express sincere gratitude to the staff of the UB Dental School, UB Anatomy Lab, & the Biomaterials Graduate
Program for supplying me with the knowledge, equipment, and motivation to accomplish this study. Special appreciation is given
to the 17 human donors of cadavers to UBs Anatomical Gifts Program.
The research methods included tensile testing, tissue-on-tissue friction testing, with HA preparations applied prophylactically or
subsequently, and HYP Assay testing. Weight and volume changes were also recorded, monitoring the effects of soaking in
DW/HA, baking, and 稗-irradiation.
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
AVGStart,1st5min
AVGStart,2nd5min
AVGStart,3rd5min
AVGStart,4th5min
AVGStart,5th5min
AVGStart,6th5min
AVG1PBS,1st5min
AVG1PBS,2nd5min
AVG1PBS,3rd5min
AVG1HA,1st5min
AVG1HA,2nd5min
AVG1HA,3rd5min
AVG1HA,4th5min
AVG1HA,5th5min
StaticFrictionCoefficient(亮)
OM Pre/Post Irrad HA/PBS (Phosphate-
Buffered Saline) Application
PreIrrad 袖 PostIrrad 袖
OM HA NonIrr
OM DW NonIrr
PC HA NonIrr
PC DW NonIrr
PC PO NonIrr
NonIrrad AVG
OM HA IrrOM DW Irr
PC DW Irr
PC HA Irr
Irr Avg
PC Avg
OM Avg
HA Avg
DW Avg
0
5
10
15
20
25
30
35
40
0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6
Stress(MPa)
Strain
Elastic Moduli of DW and HA-Soaked Samples of OM and PC,
NonIrradiated vs. Corresponding Irradiated Samples
0%
10%
20%
30%
40%
50%
60%
70%
80%
Relative%Collagen
Sample Group
AVG % Collagen](https://image.slidesharecdn.com/3c5bdea3-1a8b-4b43-a1f6-bd6396d790e9-150704162240-lva1-app6891/85/Huber_Aaron-Sigma-Xi-2015-UB-1-320.jpg)
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- 1. Influence of hyaluronan (HA) preparations on ionizing- radiation-treated collagen-based tissues Huber A.T. BS, Meyer A.E. PhD, Zhao R.G. PhD, Baier R.E. PhD Biomaterials Graduate Program, SUNY at Buffalo, Buffalo, NY, USA Abstract #__ ABSTRACT RESULTS CONCLUSIONS INTRODUCTION MATERIALS AND METHODS REFERENCES ACKNOWLEDGMENTS Objectives: It has been proposed that HA solution (.5% HA in distilled water, DW), due to its natural lubrication properties [1], could remedy xerostomic patients. Investigation was conducted using Tensile, Friction, and Hydroxyproline (HYP) Assay Studies. The experimental aim was to observe effects of 70 Gray 稗-Radiation (standard dose for head/neck cancer patients) and HA-soaking. Materials & Methods: Tensile: 19 non-irradiated specimens of fresh human oral mucosa (OM) and bovine pericardium (PC) and 35 irradiated specimens were soaked in DW or HA. Tensile testing until failure was conducted. Static Friction: 13 PC/OM samples were tested before and after 粒-irradiation, 7 DW-soaked, 6 HA-soaked. Tissues fastened to friction device, limiting slippage. HYP Assay: OM/PC samples, soaked in HA/DW, Irradiated and Non-irradiated, were tested. Developing a standard curve involved: Digestion of samples in HCl at 105 属C and adding a color reagent. Sample %Transmittance was compared to standard curve. Results: Tensile: Non-Irradiated PC/OM tissue had higher average strain (0.825 +/- 0.311 vs 0.485 +/- 0.186) and lower ultimate strength (9.858 +/- 1.528 MPa vs 13.597 +/- 4.480 MPa) than Irradiated PC/OM. PC had a slightly higher average strain of 0.726 +/- 0.124 and 13x-higher average ultimate strength of 21.818 +/- 6.398 MPa than OM. Static Friction: OM HA had about 遜 the 亮 (friction coefficient) as OM DW pre-irradiation. After irradiation, OM HA and DW 亮s merged to 0.45. PC HA had about 遜 the 亮 as PC DW before/after irradiation. HYP Assay: HCl-digestion-based HYP Assay is equally capable of hydrolyzing to their AA contents BOTH natural and irradiation- crosslinked tissues[2]. Final test will find if HCl-undigested samples will determine if irradiation had released any HYP due to collagen chain scission. Preliminary findings suggest that tissue degradation of OM is accelerated when irradiated and therefore releases more HYP when pulped, hinting at collagen chain scission being more abundant than chain crosslinking. HAs possible protection against this chain-scission is still being evaluated. Conclusion: HA solution relaxes the strain of OM. HA shields OM/PC during irradiation, which otherwise causes tissue embrittlement. HA significantly reduces friction for irradiated/non-irradiated PC/OM. Irradiation increases degradation of tissue, while increasing its elastic modulus and increasing its 亮. It has been observed that HA provides superior and reversible physical protection to collagen-rich tissues as monitored in friction testing. Considering the possible benefits[3] to irradiated head-and-neck cancer patients who have lost all natural salivary lubrication, fresh human oral mucosa was collected and tested against a crosslinked standard pericardium reference material for its ability to take up HA reversibly and preserve desirable tissue properties after exposure to clinically-used doses of 70 Gray (of Cs-137) 稗-irradiation. Adding HA relieved the tensile strain of both normal and irradiated samples, and provided some modulation of the radiation- induced crosslinking and embrittling (increase in Elastic Modulus) of OM. Weight measurements illustrated that HA was taken up into native and irradiated tissues, and was completely reversible by water exposure, so the effect was more than superficial lubrication (which lasts only short times). Therefore, the predominant mechanisms of protection of HA formulations are friction reduction in both irradiated and non-irradiated OM and strain relief, which now remain to be correlated with subjective pain relief and improved oral function. This mechanical disparity correlates with OM having 1/3 the collagen amount as PC. [1] Singh A, Corvelli M, Unterman SA, Wepasnick KA, McDonnell P, Elisseeff JH. Enhanced lubrication on tissue and biomaterial surfaces through peptide- mediated binding of hyaluronic acid. Nat Mater. 2014;13:988-95. [2] Bailey AJ, Rhodes DN, Cater CW. Irradiation-Induced Crosslinking of Collagen. Radiation Research. 1964;22:606-21. [3] Kawano T, Miura H, Mawatari T, Moro-Oka T, Nakanishi Y, Higaki H, et al. Mechanical effects of the intraarticular administration of high molecular weight hyaluronic acid plus phospholipid on synovial joint lubrication and prevention of articular cartilage degeneration in experimental osteoarthritis. Arthritis & Rheumatism. 2003;48:1923-9. I would like to express sincere gratitude to the staff of the UB Dental School, UB Anatomy Lab, & the Biomaterials Graduate Program for supplying me with the knowledge, equipment, and motivation to accomplish this study. Special appreciation is given to the 17 human donors of cadavers to UBs Anatomical Gifts Program. The research methods included tensile testing, tissue-on-tissue friction testing, with HA preparations applied prophylactically or subsequently, and HYP Assay testing. Weight and volume changes were also recorded, monitoring the effects of soaking in DW/HA, baking, and 稗-irradiation. 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 AVGStart,1st5min AVGStart,2nd5min AVGStart,3rd5min AVGStart,4th5min AVGStart,5th5min AVGStart,6th5min AVG1PBS,1st5min AVG1PBS,2nd5min AVG1PBS,3rd5min AVG1HA,1st5min AVG1HA,2nd5min AVG1HA,3rd5min AVG1HA,4th5min AVG1HA,5th5min StaticFrictionCoefficient(亮) OM Pre/Post Irrad HA/PBS (Phosphate- Buffered Saline) Application PreIrrad 袖 PostIrrad 袖 OM HA NonIrr OM DW NonIrr PC HA NonIrr PC DW NonIrr PC PO NonIrr NonIrrad AVG OM HA IrrOM DW Irr PC DW Irr PC HA Irr Irr Avg PC Avg OM Avg HA Avg DW Avg 0 5 10 15 20 25 30 35 40 0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 Stress(MPa) Strain Elastic Moduli of DW and HA-Soaked Samples of OM and PC, NonIrradiated vs. Corresponding Irradiated Samples 0% 10% 20% 30% 40% 50% 60% 70% 80% Relative%Collagen Sample Group AVG % Collagen