The aim of the present work is to investigate the effect of the He-Ne laser irradiation on the whole human blood (HB) in order to enhance the conditions of conservation. The HB was irradiated by He-Ne laser; (λ = 632 nm, continuous wave, power 30 mW, 2 mm diameter beam spot), electrical properties and complete blood count CBC were measured at three doses (0.0287, 0.0563 and 0.198 J/cm 3) to the relevant best exposure dose during storage periods 9, 24, 30, 35 & 50 days. The irradiation process with the selected dose was performed by the exposure of the laser beam on the blood sample flow through narrow tube of cross section area, 0.0831 cm 2. Blood dielectric parameters, (electric conductivity, dielectric constant, dielectric loss and dipole moment) and CBC, (red blood cell, white blood cell, hematocrit, hemoglobin, mean corpuscular volume, mean corpuscular hemoglobin, and mean cell or corpuscular hemoglobin in concentration) were measured. The obtained results were compared with that of the control and showed that the best irradiation exposure dose suitable for increasing the time of blood storage with minimum changes in properties is 0.198 J/cm 3 and storage period of about 50 days. The present study revealed that irradiation by He-Ne laser could be considered a good means to improve the conservation conditions of human blood.
Biostimulation effects of laser irradiation on living organisms were found for three decades [
The study of the low level laser irradiation effects on blood is very important in the process of revealing the mechanisms of the action of laser radiation on biological tissues, as the blood is permanently composed of a diversity of cells whose membranes contains lipids, sugars and proteins. Blood plasma probably contains the most diverse range of biological products among all tissues, some being intrinsic to other organs (amino acids, proteins, lipids, hormones, antibodies and adjusting factors), exogenous substances or component substances (clotting and defense enzymatic systems, such as complement, etc.). Some of the substances present in the blood may act as primary acceptors of radiation [
Several clinical treatments with low-power laser irradiation have been applied to various pathologic processes, such as wound healing and tissue repair, as well as remodeling [
There are widespread applications of low intensity laser irradiation in various areas of the medical field [
The most used laser of low level laser studies are He-Ne laser emitting light at a wavelength of 632.8 nm [
The blood as well as all biological tissues represents a special class of heterogeneous systems. It is a complex system where various mechanisms of polarization such as dipole, Maxwell-Wagner’s, electrochemical, etc. can be realized [
They are considered a good measure of the functional state of the membrane and cytoplasm of the cells (e.g., counter ion relaxation associated with intrinsic membrane changes, dipole reaction in cell membrane, conductivity transport in the extra-cellar medium and through the membrane and tissue water relaxation). Information about tissue structure and composition, e.g. water content or presence of a tumor, might be obtained by measuring the dielectric properties of the tissues [
The aim of the present study is to find whether as a result of low level He-Ne laser action, the possibility to increase the viability of blood and get rejuvenation, in order to perform transfusion, as well as prolong its life preserved at 4˚C.
Blood samples were obtained from 5 adult volunteers (female regular blood donors), after oral informed consent. According to local laws, neither medical ethic committee approval nor written consent is required. Blood samples from donors were referred for the standard blood analyses. In order to ensure the request of blinding method, only the coordinator of the study has access to all gathered data.
Blood samples: were prepared from the whole blood of healthy donors, preserved in Macopharma bags, and treated with CPD (sodium citrate as anticoagulant + P, D as preserver). CPD is used to store human blood for periods of about 21 days. To have more samples per volunteer, the original bags were separated into smaller bags prior to getting the blood each sample being able to collect about 10 ml of blood.
Blood bag: A Blood bag of 450 ml ± 10% (Agarry) which contains CPDA-1 was used. Most blood collection bags (adult) contain 63 ml CPDA anticoagulant Fresh human blood was obtained from ten healthydonor. Thirty small bag samples were papered to determine the normal dynamics of some dielectric, blood count parameters during the preservation period. They are divided into six groups stored, one control and five group’s irradiated with He-Ne laser. The blood samples were storage before and after irradiation in transfused bags at 6˚C - 8˚C for (9, 24, 30, 35 & 50) days. c) Irradiation method Irradiation was carried out using a source of He-Ne laser (U.S.PAT. 311, 969), continues wave, 632.8 nm, 2 mm spot diameter, 30 mW, for three doses 0.0287, 0.0563 and 0.198 J/cm3 to relevant the best exposure dose during storage periods 9, 24, 30, 35 & 50 days. The laser beam was in contact to the narrow rubber tube through which blood flow, as shown in
The dielectric measurements were carried out using LCR meter bridge (HIOKI 3531, Japan) in the frequency range 50 Hz to 100 KHz. A parallel plate conductivity cell was used with platinum electrodes, of area “A” 0.48 cm2 and separating distance “d” 1 cm. The measured parameters were capacitance c and conductance G and consequently the dielectric constant έ, the dielectric loss ε" and the conductivity σ, and electrical dipole moment D could be calculated from the following relations;
where, T is the absolute temperature, N is Avogadro’s number, K is Boltzmann’s constant and C is the hemoglobin concentration.
The plot of dielectric loss ε" against the dielectric constant ε' is in the form of a semicircle whose center lies below the abscissa and intersects the axis at the points
Since the center of the semicircle makes an angle απ/2 rad with the points (
where τ0 is the relaxation time and μ, υ are the distances on the cole-cole diagram (
The human blood (HB) was exposed to continuous waves of 30 mW He-Ne laser, 632.8 nm and 2 mm spot diameter through during preservation periods 9, 24, 30, 35, and 50 days. The obtained data of the dielectric parameters and CBC before and after irradiation are summarized in
Storage period (days) | Δε × 106 | α | τ × 10−6 | τ (cole) × 10−6 | D × 10−13 | fs (Hz) | ||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Control | Irrad. | Control | Irrad. | Control | Irrad. | Control | Irrad. | Control | Irrad. | Control | Irrad. | |
9 | 204 | 206 | 0.119 | 0.106 | 505.9 | 482 | 337.26 | 321.3 | 4.18 | 4.205 | 307.5 | 330.19 |
24 | 212 | 196 | 0.113 | 0.101 | 518.28 | 537.6 | 345.59 | 358.4 | 4.26 | 4.102 | 307.08 | 296.04 |
30 | 206 | 154 | 0.107 | 0.0947 | 536.76 | 546.1 | 357.33 | 364.1 | 4.20 | 3.636 | 296.5 | 291.4 |
35 | 204 | 204 | 0.101 | 0.0885 | 665.9 | 563 | 443.9 | 375.33 | 4.18 | 4.184 | 238.97 | 282.6 |
50 | 208 | 210 | 0.0947 | 0.0822 | 680.5 | 641.8 | 453.66 | 427.86 | 4.22 | 4.245 | 233.8 | 247 |
The changes in HB conductivity due to laser irradiation dose of 0.198 J/cm3 during storage periods 9, 24, 30, 35 and 50 days is shown in
The conductivity of irradiated HB samples showed the same behavior of change with frequency as that of control samples but with lower values for all days of storage. This drop in conductivity may be attributed to the energy obtained by hemoglobin and oxygen activation and also due to laser irradiation which in turn lead to an increase in the cell membrane resistance which become more protective with increasing the storage period.
The relation between the dielectric loss (ε'') and dielectric constant (έ) of the human blood , in the frequency range from 50 Hz to 100 kHz is shown in
It is clear from these figures that cole-cole plot for all samples were semicircles in which the control days are lower than that the irradiated .This is due to the increase in dielectric constant έ and dielectric loss ε" values after irradiation in comparison with control values. The increase in έ is perhaps due to higher polarization of ions in RBCs and water, while the increase of dielectric loss is due to the increase in dipole moment after irradiation as shown in
The center of the semicircles is not located at the x-axis, but deviated below that axis where the radius of the circle making an angle (απ/2) with the έ axis. Tan απ/2 is determined and hence the phase angle (α) is calculated. The experimental data of the variation in dielectric loss ε" of HB with permittivity έ for control sample (without irradiation) and irradiated samples during preservation days and other dielectric parameters calculated from the last figures are summarized in
The blood was collected in bags with anticoagulant and preserving solution undergoes, through conservation days (9, 24, 30, 35, and 50) at 4˚C to 6˚C, a natural process of aging or destruction because of stress. The CBC is
favored process for getting the information about the functional state of the blood during storages days for control and irradiated samples as shown in
The complete blood count CBC, represented the parameters, (red blood cell, RBC, white blood cell, WBC, hematocrit, HCT, mean corpuscular volume, MCV, mean corpuscular hemoglobin, MCH, and corpuscular hemoglobin in concentration, MCHC, as in
The increase in Δε of RBC after irradiation is due to cell membrane activation of laser stimulus, which in turn cause diffusion of K+, Na+ ions and absorption of laser energy by hemoglobin, these leading that the Na-K pump performed to recovery of cell membrane, while the increase in WBC in control samples perhaps due to defense of this cells for any abnormal variations in blood as the cell membrane of RBC is damage or its hemolysis while the decrease of WBC after irradiated during preservation refer to recovery of cell membrane and excess of protective of blood cell produced from the effects He-Ne laser at dose 0.198 J/cm3.
The increase in hematocrit HCT is due the increase in RBCs [
The values of MCV after irradiated slowly decreases than control values. Also MCH and MCHC values were slowly increases than control as shown in
Days | RBCs | WBC | HCT (%) | MCV (μm3) | MCH (pg) | MCHC (%) | ||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Control | Irrad | Control | Irrad | Control | Irrad | Control | Irrad | Control | Irrad | Control | Irrad | |
9 | 3.84 | 4.01 | 4.5 | 4.6 | 34.2 | 34.4 | 86.7 | 85.9 | 30.2 | 29.9 | 34.8 | 34.9 |
24 | 4.13 | 4.21 | 7.1 | 3.1 | 34.5 | 35.4 | 86.5 | 84.1 | 28.8 | 29.5 | 34.5 | 35 |
30 | 3.67 | 4 | 8.8 | 5.5 | 31.4 | 34.6 | 85.5 | 86.6 | 30 | 30.3 | 35 | 35 |
35 | 4.01 | 4.08 | 9.2 | 5.1 | 33.2 | 35.2 | 87.5 | 88.5 | 30.5 | 30.6 | 35.1 | 35.1 |
50 | 4.1 | 4.21 | 10.1 | 4.7 | 37.1 | 38.1 | 90.5 | 90.4 | 32 | 32.1 | 35.3 | 35.4 |
slight trend toward decrease and the Na+ ions increase for certain dose (0.2 J/cm3) agree with [
The total conductivity (σ) of materials depends, in general, on its chemical composition, and hence the living tissues conductivity may also be affected by the changes in their chemical composition is very important because it may reflect the health status of the human.
From the obtained results it is evident that, the high value of the control conductivity (σ) during storage days may be attributed to the high value of the membrane capacitance and conductance due to roughly explicable considering that the microvillus structure allows an higher ionic permeation with an increase of the ionic hydration and also due to the inner solution (ionic hemoglobin) in RBCs [
The results of CBC analysis support the dielectric parameters measurements, the conductivity of HB after irradiation decreases than its control value as in
The dielectric loss curve can be evaluated by calculating its total area. It is proportional to the total concentration of dipoles in blood. The values of dipole moment for hemoglobin are registered in table [
In this study we had investigated the variation in HB dielectric parameters (σ, ε', ε'', τ & D), due to laser irradiation as indication on the membrane structure and molecular motion. We mainly followed the conductivity variation, corroborated with polarization of ε', ε'' and dipole moment D. Also, this study investigates the variation in CBC parameters; RBC, WBC, HCT, MCV, MCH and MCHC, for irradiated and non-irradiated blood, as a function of preservation periods. In case of CBC parameters we mainly followed MCV, variation, corroborated with MCHC, MCH.
Generally, He-Ne laser radiation acts in the direction of maintaining the shape of cells, without leading to negative effect as spherocytosis and hemolysis, which normally occur in non-irradiated blood during the preservation period.
The obtained data suggested that most of the parameters under measurement were significantly modified by low level He-Ne laser effects with action of dose 0.198 J/cm3, and consequently positively influenced on the preserved days.
The present study revealed that irradiation by He-Ne laser could be considered a good means to improve the conservation conditions of human blood.
Samira M.Sallam,Abdelsattar M.Sallam,El-Sayed M.El-Sayed,L. I. AboSalem,Mona M.Rizk, (2015) Enhancement of Human Blood Storage Period by Irradiation of Low Level He-Ne Laser. Journal of Biophysical Chemistry,06,77-86. doi: 10.4236/jbpc.2015.63008