Effect of Humid Air Exposed to IR Radiation on Enzyme Activity.

Water vapor absorbs well in the infra-red region of the electromagnetic spectrum. Absorption of radiant energy by water or water droplets leads to formation of exclusion zone water that possesses peculiar physico-chemical properties. In the course of this study, normally functioning and damaged alkaline phosphatase, horseradish peroxidase and catalase were treated with humid air irradiated with infrared light with a wavelength in the range of 1270 nm and referred to as coherent humidity (CoHu). One-minute long treatment with CoHu helped to partially protect enzymes from heat inactivation, mixed function oxidation, and loss of activity due to partial unfolding. Authors suggest that a possible mechanism underlying the observed effects involves altering the physicochemical properties of aqueous media while treatment of the objects with CoHu where CoHu acts as an intermediary.

Fosfatasa alcalina ósea sérica elevada no mediada por metástasis en un paciente con cáncer renal / Elevated bone alkaline phosphatase not mediated by metastasis in a patient with renal cancer

Presentamos un paciente de 63 años con cáncer renal y aumento de fosfatasa alcalina sérica de tipo óseo de acuerdo con su reactividad con anticuerpos monoclonales específicos. Se descartaron las causas conocidas de aumento de la isoenzima, incluyendo metástasis óseas. Los niveles enzimáticos cayeron abruptamente con la remoción del tumor, por lo que consideramos a este último como su origen. Diversas isoenzimas de fosfatasa alcalina pueden ser producidas y secretadas por tumores como manifestación paraneoplásica. El conocimiento de esto puede, en ocasiones, orientarnos hacia la presencia de una neoplasia oculta. Además, los cambios en los niveles séricos de esas isoenzimas pueden ser indicadores de respuesta al tratamiento o de recidiva tumoral. (AU)

A 63-year old man was seen in the outpatient clinic because of renal cancer and elevation in bone alkaline phosphatase measured by monoclonal antibodies assay. Known causes of bone isoenzyme augmentation, including bone metastases, were ruled out. The tumoral origin of the isoenzyme was diagnosed because after removal of the tumor the enzymatic levels fell sharply. Several alkaline phosphatase isoenzymes can be produced and secreted by tumors as a paraneoplasic manifestation and their elevation could be a manifestation of an occult neoplasia. Furthermore the monitoring of their blood levels can be useful means of treatment response and a tool to monitoring recurrence if a sharp decrease after removal of the tumor is observed. (AU)

Low level laser therapy modulates viability, alkaline phosphatase and matrix metalloproteinase-2 activities of osteoblasts.

Low level laser therapy (LLLT) has been shown to stimulate bone cell metabolism but their impact on the matrix metalloproteinase (MMP) expression and activity is little explored. This study evaluated the influence of LLLT at two different wavelengths, red and infrared, on MC3T3-E1 preosteoblast viability, alkaline phosphatase (ALP) and MMP-2 and -9 activities. To accomplish this, MC3T3-E1 cells were irradiated with a punctual application of either red (660nm; InGaAIP active medium) or infrared (780nm; GaAlAs active medium) lasers both at a potency of 20mW, energy dose of 0.08 or 0.16J, and energy density of 1.9J/cm2 or 3.8J/cm2, respectively. The control group received no irradiation. Cellular viability, ALP and MMP-2 and -9 activities were assessed by MTT assay, enzymatic activity and zymography, respectively, at 24, 48 and 72h. The treatment of cells with both red and infrared lasers significantly increased the cellular viability compared to the non-irradiated control group at 24 and 48h. The ALP activity was also up modulated in infrared groups at 24 and 72h, depending on the energy densities. In addition, the irradiation with red laser at the energy density of 1.9J/cm2 promoted an enhancement of MMP-2 activity at 48 and 72h. However, no differences were observed for the MMP-9 activity. In conclusion, when used at these specific parameters, LLL modulates both preosteoblast viability and differentiation highlighted by the increased ALP and MMP-2 activities induced by irradiation.

[Genic and non-genic regulation of low frequence pulsed electromagnetic fields on osteoblasts differentiation].

Objective: To study the genic and non-genic regulation of 50 Hz 0.6 mT pulsed electromagnenic fields (PEMF) on rat calvarial osteoblasts (ROB) differentiation. Methods: ROBs were achieved by enzyme digestion, and treated with 50 Hz 0.6 mT PEMFs for 1.5 hours after subculture. The alkaline phosphatase (ALP) activity, mRNA transcription of ALP, Runx2 and OSX and protein expression of Runx2 and OSX were detected at 0, 3, 6, 9 and 12 hours after PEMF treatment. Results: The ALP activity at 3 hours after treatment was significantly higher than that in the control(P<0.01), while the mRNA transcription of ALP began to increase at 6 hours after treatment. The mRNA transcription of Runx2 increased immediately after treatment and regressed at 6 hours, then increased again. The protein expression of it corresponded but with a little lag. The mRNA transcription of OSX also raised instantly after treatment, then returned to the level of control at 6 hours, and lower than control at 12 hours significantly. The protein expression of it also corresponded but with a bit delay. Conclusions: There are genic regulation for the protein expression of Runx2 and OSX, and non-genic regulation for the ALP activity on the process of 50 Hz 0.6 mT PEMFs prompts ROBs differentiation.

Phototherapy up-regulates dentin matrix proteins expression and synthesis by stem cells from human-exfoliated deciduous teeth.

OBJECTIVES: The aim of this study was to evaluate the effects of infrared LED (850nm) irradiation on dentin matrix proteins expression and synthesis by cultured stem cells from human exfoliated deciduous teeth (SHED). METHODS: Near-exfoliation primary teeth were extracted (n=3), and SHED cultures were characterized by immunofluorescence using STRO-1, CD44, CD146, Nanog and OCT3/4 antibodies, before experimental protocol. The SHEDs were seeded (3×10(4) cells/cm(2)) with DMEM containing 10% FBS. After 24-h incubation, the culture medium was replaced by osteogenic differentiation medium, and the cells were irradiated with LED light at energy densities (EDs) of 0 (control), 2, or 4J/cm(2) (n=8). The irradiated SHEDs were then evaluated for alkaline phosphatase (ALP) activity, total protein (TP) production, and collagen synthesis (SIRCOL™ Assay), as well as ALP, collagen type I (Col I), dentin sialophosphoprotein (DSPP), and dentin matrix acidic phosphoprotein (DMP-1) gene expression (qPCR). Data were analyzed by Kruskal-Wallis and Mann-Whitney tests (α=0.05). RESULTS: Increased ALP activity and collagen synthesis, as well as gene expression of DSPP and ALP, were observed for both EDs compared with non-irradiated cells. The ED of 4J/cm(2) also increased gene expression of COL I and DMP-1. CONCLUSIONS: In conclusion, infrared LED irradiation was capable of biostimulating SHEDs by increasing the expression and synthesis of proteins related with mineralized tissue formation, with overall better results for the energy dose of 4J/cm(2). CLINICAL SIGNIFICANCE: Phototherapy is an additional approach for the clinical application of LED in Restorative Dentistry. Infrared LED irradiation of the cavity's floor could biostimulate subjacent pulp cells, improving local tissue healing.

Effects of 10-GHz microwaves on hematological parameters in Swiss albino mice and their modulation by Prunus avium.

The objective of this study was to investigate the modulatory role of Prunus avium fruit extract (PAE) on several blood parameters after exposure to 10-GHz microwaves. Swiss albino mice from an inbred colony were selected and divided into 3 groups. Mice in group I served as the control; they were placed in a Plexiglas cage (without energizing the system) for 2 hours/day for 30 consecutive days. Group II mice were exposed to 10-GHz microwaves for 2 hours/day for 30 consecutive days. Mice in group III received PAE (500 mg/kg/body weight) orally once daily 1 hour before exposure to 10-GHz microwaves (2 hours/day) for 30 consecutive days. After 30 days of treatment, blood samples were collected from mice in all groups and analyzed. Hemoglobin, monocytes, packed cell volume, red blood cells, mean corpuscular hemoglobin concentration declined significantly (P ≤ 0.01), whereas white blood cells, lymphocytes, erythrocyte sedimentation rate, and mean corpuscular volume increased significantly (P ≤ 0.01) compared to the control group (group I). Cholesterol, alkaline phosphatase, and lipid peroxidation also increased significantly (P ≤ 0.01). Depletion in blood sugar, total protein, acid phosphatase, and glutathione levels was noted after microwave exposure compared with levels in the sham-exposed (control) mice. Histopathological alterations in blood cells also were seen. Signs of improvements in the hematological, biochemical, and histopathological parameters were recorded in group III, where PAE was supplemented before exposure. Exposure to microwaves influences hematological parameters, which could be ameliorated by the supplementation of PAE.

Low-power laser irradiation promotes the proliferation and osteogenic differentiation of human periodontal ligament cells via cyclic adenosine monophosphate.

Retaining or improving periodontal ligament (PDL) function is crucial for restoring periodontal defects. The aim of this study was to evaluate the physiological effects of low-power laser irradiation (LPLI) on the proliferation and osteogenic differentiation of human PDL (hPDL) cells. Cultured hPDL cells were irradiated (660 nm) daily with doses of 0, 1, 2 or 4 J⋅cm(-2). Cell proliferation was evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and the effect of LPLI on osteogenic differentiation was assessed by Alizarin Red S staining and alkaline phosphatase (ALP) activity. Additionally, osteogenic marker gene expression was confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR). Our data showed that LPLI at a dose of 2 J⋅cm(-2) significantly promoted hPDL cell proliferation at days 3 and 5. In addition, LPLI at energy doses of 2 and 4 J⋅cm(-2) showed potential osteogenic capacity, as it stimulated ALP activity, calcium deposition, and osteogenic gene expression. We also showed that cyclic adenosine monophosphate (cAMP) is a critical regulator of the LPLI-mediated effects on hPDL cells. This study shows that LPLI can promote the proliferation and osteogenic differentiation of hPDL cells. These results suggest the potential use of LPLI in clinical applications for periodontal tissue regeneration.

Effect of radiation on the Notch signaling pathway in osteoblasts.

Notch signaling has been shown to be important in osteoblast differentiation. Therapeutic radiation has been shown to alter the skeletal system, yet little information is available on the changes in Notch signaling in irradiated osteoblasts. The purpose of this study was to analyze the effect of radiation therapy with 2 and 4 Gy on Notch signaling in osteoblasts. In order to assess the radiation damage on osteoblast differentiation, total RNA and protein were collected three days after exposure to radiation. The effects of radiation on Notch signaling at the early and terminal stages of osteoblastic MC3T3-E1 cell differentiation was analyzed by qRT-PCR and western blot analysis. Our study applied a previously established method to induce MC3T3-E1 cell differentiation into osteoblasts and osteoblast precursors. Our results showed that the expression of Notch receptors (Notch1-4), ligands (Jagged1, Jagged2 and Delta1), target of Notch signaling (Hes1) and markers (ALP, M-CSF, RANKL and OPG) were altered following 2 and 4 Gy of irradiation. The present research did not indicate a strong relationship between Notch1 regulation and suppression of osteoblast differentiation. We found Hes1 may play a role in the radiation effect on osteoblast differentiation. Our results indicate that radiated osteoblast precursors and osteoblasts promoted osteoclast differentiation and proliferation.

Effect of low level laser therapy and zoledronate on the viability and ALP activity of Saos-2 cells.

A limited number of clinical studies indicate the supportive role of low level laser therapy (LLLT) on medical and/or surgical approaches carried out in treatment modalities for bisphosphonate related necrosis of jaws (BRONJ), the most common side effect of bisphosphonates used to inhibit bone resorption. The purpose of this study was to investigate the effects of LLLT on cell proliferation and alkaline phosphatase (ALP) activity of human osteoblast-like cells (Saos-2) treated with different doses of zoledronate, the most potent bisphosphonate. Saos-2 cells were treated with different concentrations of zoledronate and were irradiated with diode laser (wavelength 808 nm, 10 s, 0.25 or 0.50 W). Cell numbers and ALP activity of the cells were determined. LLLT mildly increased the proliferation rate or ALP activity, while zoledronate reduced both. When applied together, LLLT lessened the detrimental effects of zoledronate and improved cell function and/or proliferation. Based on the results of this study, it was concluded that LLLT has biostimulative effects on Saos-2 cells, even after treatment with zoledronate. LLLT may serve as a useful supportive method for BRONJ treatment through enhancement of healing by osteoblasts.

Effects of sterilization and storage on the properties of ALP-grafted biomaterials for prosthetic and bone tissue engineering applications.

Grafting of the biomaterial surfaces with biomolecules is nowadays a challenging research field for prosthetic and bone tissue engineering applications. On the other hand, very few research works investigate the effect of the sterilization processes on the properties of functionalized biomaterials. In this study, the effects of different sterilization techniques (e.g. gamma and electron beam irradiation, ethylene oxide) on the enzymatic activity of bioactive glasses and Ti6Al4V grafted with alkaline phosphatase (ALP) have been analyzed. Sterility maintenance and in vitro bioactivity of the sterilized surfaces have also been investigated. Finally the effect of packaging and storage conditions has been considered.

In vitro effects of low frequency electromagnetic fields on osteoblast proliferation and maturation in an inflammatory environment.

An in vitro model was set up to investigate the effects of low frequency pulsed electromagnetic fields (PEMF) and its induced electric fields on osteoblast cells under inflammatory conditions. Osteoblasts (7F2) were seeded on top of chitosan scaffolds and co-cultured with macrophage cells (RAW 264.7) growing on the bottom of culture wells, stimulated by lipopolysaccharide to release reactive oxygen species including nitric oxide (NO). The co-culture was exposed to PEMF (magnitude of the magnetic field = 1.5 mT; induced electric voltage = 2.5 mV; frequency = 75 Hz; pulse duration = 1.3 ms) for 9 h. The osteoblasts were examined for their proliferation, viability, alkaline phosphatase (ALP) activity, and genetic expressions of type I collagen (COL I) and osteocalcin (OC), immediately and 7 days after PEMF exposure (days 0 and 7). Macrophage cell viability and NO concentration in the medium were monitored before and after PEMF exposure. The PEMF-exposed co-culture released a significantly higher amount of NO (65 µM) compared to control (17 µM) on day 7. Despite the high level of NO in the medium that was reported to be cytotoxic, PEMF-exposed osteoblasts had enhanced cell proliferation (23%), viability (36%), and COL I mRNA expression (3.4-fold) compared to the controls. The osteoblasts subjected to the PEMF had 41% less ALP activity than the control, which was associated with the active cell proliferation and COL I expression. The expression of OC mRNA was not seen in either the PEMF or control group, indicating cells had not entered the mineralization stage by day 7.

High power-pulsed Nd:YAG laser as a new stimulus to induce BMP-2 expression in MC3T3-E1 osteoblasts.

BACKGROUND AND OBJECTIVE: High-power laser has recently become a physical stimulus for bone regeneration. Little is known about how high-power laser irradiation affects osteoblast differentiation. This study investigated osteoblast responses to high-power laser and combined irradiation with BMP-2 treatment. STUDY DESIGN/MATERIALS AND METHODS: MC3T3-E1 pre-osteoblasts were exposed to laser irradiation, 100 ng/ml BMP-2 or both. Cells were irradiated with a Q-switched, pulsed neodymium-doped yttrium aluminum garnet (Nd:YAG) laser, with a 1,064 nm wavelength and 0.75 W output power under 1.5, 3, or 5 J/cm(2) energy densities. Cell proliferation was evaluated using tetrazolium salt, WST-8. To determine the effect of these treatments on in vitro osteogenesis, we examined alkaline phosphatase (ALP) activity, mineral deposition, and expression of genes associated with osteogenesis. Quantitative real time PCR or ELISA was used to examine cytokine expression. In each experiment, either non-irradiated or BMP-2 (100 ng/ml)-treated cells were used as controls. RESULTS: High-power, low-level, Nd:YAG laser irradiation significantly increased ALP activity, when combined with BMP-2 or not. Cell proliferation declined in the irradiation and combined irradiation/BMP-2 groups. Interestingly, Nd:YAG laser stimulation resulted in significant induction of endogenous BMP-2 protein and gene expression. The increased expression of upstream regulators cbfa1 by Nd:YAG laser alone was comparable to exogenous BMP-2 treatment (100 ng/ml). Combined laser/BMP-2 treatment was synergistic in the expression of some genes (IGF-1, cbfa1) and ALP activity, compared to both BMP-2 treatment and laser irradiation alone. In vitro matrix mineralization was significantly accelerated by laser stimulation compared to that of the control, more so than with the combined laser/BMP-2 treatment. CONCLUSIONS: The present in vitro findings demonstrate that high-power, low-level Nd:YAG laser increased osteoblast activity, very efficiently accelerating mineral deposition. Osteoinductive effect of laser is likely mediated by activation of BMP-2-related signaling pathway.

Effects of repeated extracorporeal shock wave in urinary biochemical markers of rats.

PURPOSE: To access the effect of repeated extracorporeal shock wave (ESW) on urinary biochemical markers METHODS: 20 rats were assigned for ESW (Direx Tripter X1(R) - 14 KV) to one of two groups: G1 (n=10) one ESW; G2 (n=10) two ESWs within a 14-day interval. Within the twenty-four hour period before and after the application of shock waves, the animals were placed in metabolic cages for 24 hour urine collection. The ph, creatinine, sodium, potassium, chlorides, calcium, magnesium, phosphorus, oxalates, alkaline phosphatase and citrates were measured. Twenty-four hours after the material was collected for urinary determination, the animals underwent nephrectomy of the kidney submitted to the ESW applications and were, then, sacrificed. The kidneys were processed for histopathological examination. RESULTS: Small variations in the biochemical markers were found in both groups, with no significant differences between the values obtained either prior to or following the ESW applications, except for citrate and alkaline phosphatase. Citraturia decreased significantly in group 2, following the second ESWL application (24.8 +/- 3.0 mg/day after the first ESWL vs. 15.3 +/- 2.2 mg/day after the second ESWL; p < 0.05). Alkaline phosphatase increased significantly following ESWL in group I (0.57 +/- 0.02 vs. 0.79 +/- 0.04 micromol/mg creatinine; p < 0.01) and also in group 2 (0.69 +/- 0.05 vs. 0.83 +/- 0.03 micromol/mg creatinine; p < 0.05). Glomerular, interstitial and sub-capsular hemorrhage with perivascular edema was found in the animals in both groups studied. CONCLUSIONS: A significant increase in urinary alkaline phosphatase was found in both groups studied, suggesting a proximal tubule lesion. In the group of rats undergoing more than one ESWL application, a smaller urinary citrate excretion was noticed, which may be a factor contributing for the formation of new calculi.

Effects of repeated extracorporeal shock wave in urinary biochemical markers of rats / Avaliação dos fatores bioquímicos urinários de risco para nefrolitíase em ratos submetidos à aplicação repetida de ondas de choque eletro-hidráulicas

PURPOSE: To access the effect of repeated extracorporeal shock wave (ESW) on urinary biochemical markers METHODS: 20 rats were assigned for ESW (Direx Tripter X1® - 14 KV) to one of two groups: G1 (n=10) one ESW; G2 (n=10) two ESWs within a 14-day interval. Within the twenty-four hour period before and after the application of shock waves, the animals were placed in metabolic cages for 24 hour urine collection. The ph, creatinine, sodium, potassium, chlorides, calcium, magnesium, phosphorus, oxalates, alkaline phosphatase and citrates were measured. Twenty-four hours after the material was collected for urinary determination, the animals underwent nephrectomy of the kidney submitted to the ESW applications and were, then, sacrificed. The kidneys were processed for hispatological examination. RESULTS: Small variations in the biochemical markers were found in both groups, with no significant differences between the values obtained either prior to or following the ESW applications, except for citrate and alkaline phosphatase. Citraturia decreased significantly in group 2, following the second ESWL application (24.8 ± 3.0 mg/day after the first ESWL vs. 15.3 ± 2.2 mg/day after the second ESWL; p < 0.05). Alkaline phosphatase increased significantly following ESWL in group I (0.57 ± 0.02 vs. 0.79 ± 0.04 µmol/mg creatinine; p < 0.01) and also in group 2 (0.69 ± 0.05 vs. 0.83 ± 0.03 µmol/mg creatinine; p < 0.05). Glomerular, interstitial and sub-capsular hemorrhage with perivascular edema was found in the animals in both groups studied. CONCLUSIONS: A significant increase in urinary alkaline phosphatase was found in both groups studied, suggesting a proximal tubule lesion. In the group of rats undergoing more than one ESWL application, a smaller urinary citrate excretion was noticed, which may be a factor contributing for the formation of new calculi.

OBJETIVO: Avaliar os efeitos renais das ondas de choque eletro-hidráulicas (OCEH), utilizando como parâmetros marcadores bioquímicos urinários. MÉTODOS: Foram utilizados 20 ratos machos, EPM - Wistar, distribuídos aleatoriamente em dois grupos: G1 (n=10) Animais submetidos a uma sessão de OCEH. G2 (n=10) Animais submetidos a duas sessões de OCEH separadas por um intervalo de 14 dias. Para coleta da urina os animais foram mantidos em gaiolas metabólicas 24 horas antes e depois da aplicação das OCEH. Foram medidos o pH, a creatinina, sódio, potássio, cloretos, cálcio, magnésio, fósforo, oxalato, fosfatase alcalina e citrato. Vinte e quatro horas após a coleta da urina os animais foram submetidos à nefrectomia do rim envolvido no experimento e, em seguida, sacrificados. Os rins foram então submetidos aos procedimentos de fixação e coloração histológica com hematoxilina e eosina. RESULTADOS: Pequenas variações nos marcadores bioquímicos foram detectadas nos dois grupos, sem diferenças significantes nos valores obtidos antes e após a aplicação das OCEH, exceto para os valores urinários de citrato e fosfatase alcalina. A citratúria diminuiu significantemente nos animais do Grupo 2 após a segunda aplicação das OCEH( 24,8 ± 3,0 mg/dia após a primeira sessão de OCEH e 15,3 ± 2,2 mg/dia após a segunda sessão de OCEH; p < 0.05). A fosfatase alcalina urinária aumentou de forma significante no grupo 1 após a sessão de OCEH (0,57 ± 0,02 vs. 0,79 ± 0,04 µmol/mg de creatinina; p < 0,01) e também no grupo 2 (0,69 ± 0,05 vs. 0,83 ± 0,03 µmol/mg de creatinina; p < 0,05). Os achados histológicos observados nos animais dos dois grupos foram: hemorragia glomerular, intersticial e subcapsular, acompanhada de edema perivascular. CONCLUSÕES: Observou-se um aumento significante da fosfatase alcalina urinária nos dois grupos estudados, sugerindo uma lesão dos túbulos proximais causada pelas ondas de choque eletro-hidráulicas. Nos animais submetidos a mais de uma ...

Protection against radiation-induced testicular damage in Swiss albino mice by Mentha piperita (Linn.).

The protective effects of Mentha piperita leaf extract against radiation-induced damage in testis of Swiss albino mice have been studied. Animals (Male Swiss albino mice) were given M. piperita leaf extract orally (1 g/kg body weight/day) for three consecutive days before radiation exposure (8 Gy gamma-radiation). Mice were autopsied at 1, 3, 7, 14, and 30 days after irradiation to evaluate the radiomodulatory effect in terms of histological alterations, lipid peroxidation, and acid and alkaline phosphatases levels in testis. Radiation treatment showed reduction in the testis weight during all days of observation, however, in the M. piperita leaf extract-pretreated irradiated group there was a significant increase in testis weight. Radiation treatment induced moderate to severe testicular atrophy with degeneration of germ cells in seminiferous tubules. The tubules were shrunken and greatly depleted of germ cells. Sertoli cells with few germ cells were observed in the lumen. However, animals pre-treated with M. piperita leaf extract and exposed to radiation showed normal testicular morphology with regular arrangement of germ cells and slight degeneration of seminiferous epithelium. Significant decreases in the lipid peroxidation and acid phosphatase level and increase in level of alkaline phosphatase were observed in testis. The M. piperita leaf extract showed high amount of phenolic content, flavonoids content and flavonols. The results of the present study suggest that M. piperita has a significant radioprotective effect and the amount of phenolic compounds, the content of flavonoids and flavonols of M. piperita leaf extract may be held responsible for radioprotective effect due to their antioxidant and radical scavenging activity.

Effects of 100 GHz radiation on alkaline phosphatase activity and antigen-antibody interaction.

Equipment that generates microwave radiation (MWR) spanning the frequency range of 300 MHz-100 GHz is becoming more common. While MWR lacks sufficient energy to break chemical bonds, the disagreement as to whether MWR exposure is detrimental to cellular dysfunction may be difficult to clarify using complex systems such as whole animals, cells, or cell extracts. Recently, the high frequency range of terahertz (THz) radiation has been explored and sources of radiation and its detectors have been developed. THz radiation is associated with the frequency interval from 100 GHz to 20 THz and constitutes the next frontier in imaging science and technology. In the present study, we investigated the effect of radiation in the low frequency THz range (100 GHz) on two defined molecular interactions. First, the interaction of soluble or immobilized calf alkaline phosphatase with the substrate p-nitrophenylphosphate and second, the interaction between an antibody (mouse monoclonal anti-DNP) and its antigen (DNP). Irradiation of enzyme either prior to addition of substrate or during the enzymatic reaction resulted in small but significant reductions in enzyme activity. These differences were not observed if the enzyme had previously been immobilized onto plastic microwells. Exposure of immobilized antigen to radiation did not influence the ability of the antigen to interact with antibody. However, irradiation appeared to decrease the stability of previously formed antigen-antibody complexes. Our data suggest that 100 GHz radiation can induce small but statistically significant alterations in the characteristics of these two types of biomolecular interactions.

Effects of Smads and BMPs induced by Ga-Al-As laser irradiation on calcification ability of human dental pulp cells.

We investigated the effects of Ga-Al-As laser irradiation on the mineralization ability of human dental pulp (HDP) cells and on Smads and bone morphogenetic protein (BMP) production as one mechanism for the transmission of laser photochemical energy to cells. HDP cells in vitro were irradiated once with a Ga-Al-As laser at 1.0 W for 500 s, and calcified nodule formation was assessed by Alizarin red S staining. The laser irradiation was greater in the laser-irradiated group than in the non-irradiated group. Both calcium production and alkaline phosphatase (ALP) activity were higher after laser irradiation. Expression of mRNAs for Smad1, Smad7, BMPs, ALP, and osteocalcin was greater after laser irradiation, whereas expression of Smad6 mRNA was inhibited. Production of BMP-2 and BMP-4 in conditioned medium was also higher after laser irradiation. These results suggest that Smads and BMPs play important roles in ALP activity and calcification upon laser irradiation of HDP cells.

Relevance of laser irradiance threshold in the induction of alkaline phosphatase in human osteoblast cultures.

Induction of matrix synthesis by low-level laser has been demonstrated extensively. However, the question of dose- or power intensity-dependency is under-investigated. To address this issue we chose human osteoblast cell cultures and measured their alkaline phosphatase (ALP) activity after laser irradiation. The cell cultures were irradiated periodically by 690 nm radiation via optical transmission fiber-based laser needles, reaching into the culture dishes. The osteoblasts showed no induction of ALP activity when we used a single laser needle stimulation with a laser irradiance of 51 mW/cm(2), an increase of approximately 43% at 102 mW/cm(2) irradiance (two needles per well) and a ninefold increase at 204 mW/cm(2) irradiance (four needles per well), leaving the temperature of the culture medium unaffected. We concluded that the osteoblastic response in ALP activity to a laser stimulus shows a logarithmic relationship, with a distinct threshold, rather than a linear dose-dependency. Secondly, the laser irradiance, rather than the dose, is relevant for the impact of the laser.

Effects of gamma irradiation on mechanical behavior and calcification of glutaraldehyde-fixed bovine pericardium.

OBJECTIVE: To evaluate the effect of gamma irradiation on glutaraldehyde-fixed bovine pericardium. METHOD: Glutaraldehyde-fixed bovine pericardium was exposed to gamma radiation (doses from 0 to 10000 Gy). Six samples from each of nine groups were evaluated by optic microscopy, and shrinking and mechanical tests and the denaturation temperature was determined. Additionally, they were subcutaneously implanted in rats and after four months they were explanted and Ca2+ levels measured by atomic absorption spectroscopy. RESULTS: The Ca2+ levels were (in microg/mg): control (0 Gy) - 194.45; 50 Gy - 154.64; 100 Gy - 169.37; 200 Gy - 163.64; 500 Gy - 199.89; 1000 Gy - 184.02; 2000 Gy - 198.95; 5000 Gy - 227.95; 10000 Gy - 362.62. Gamma irradiation caused a significant effect on the biomechanical properties of the tissue. CONCLUSION: e-fixed bovine pericardium.

Enzymatic activity of alkaline phosphatase inside protein and polymer structures fabricated via multiphoton excitation.

We demonstrate micron scale control of bioactivity through the use of multiphoton excited photochemistry, where this technique has been used to cross-link three-dimensional matrixes of alkaline phosphatase, bovine serum albumin, and polyacrylamide and combinations therein. Using a fluorescence-based assay (ELF-97), the enzymatic activity has been studied using a Michaelis-Menten analysis, and we have measured the specificity constants kcat/KM for alkaline phosphatase in both the protein and polymer matrixes to be on the order of 10(5)-10(6) M(-1) s(-1)and are comparable to known literature values in other environments. It is found that the enzyme is simply entrapped in the polymer matrix, whereas it is completely covalently bound in the protein structures. The relative reaction rate of alkaline phosphatase bound to BSA with the ELF substrate was measured as a function of cross-link density and was found to decrease in the more tightly formed matrixes, indicating a decrease in the diffusion in the matrix.