Search

Planar MFC prototypes were constructed and experimented to operate as sensors of the anoxic condition in a denitrification tank of a wastewater treatment plant in Italy, during different times in 2018 – 2019. Electrodes were differently enriched with carbon paint containing nanotubes and CeO2 nanoparticles. Performances of different electrodes were compared. Results underline critical anoxic conditions in the tank, that caused a very low signal and phenomena of signal reversion during some period of the year. the activity of aerobic microorganisms and protozoa growing and grazing the bacteria on the electrodes strongly influenced the signal of the MFCs. The presence of nanoceria enhanced, for some extent, the MFC signal, both in presence of reversing trends and in absence of these phenomena. In absence of reversing trends, nanoceria enhanced the MFC voltage. Such signal trends from MFCs can give, in real-time, useful information to optimize the purification process without the necessity of frequent biological and chemical analyses.

Neonatal brain inflammation produced by intraperitoneal (i.p.) injection of lipopolysaccharide (LPS) results in long-lasting brain dopaminergic injury and motor disturbances in adult rats. The goal of the present work is to investigate the effect of neonatal systemic LPS exposure (1 or 2 mg/kg, i.p. injection in postnatal day 5, P5, male rats)-induced dopaminergic injury to examine methamphetamine (METH)-induced behavioral sensitization as an indicator of drug addiction. On P70, subjects underwent a treatment schedule of 5 once daily subcutaneous (s.c.) administrations of METH (0.5 mg/kg) (P70-P74) to induce behavioral sensitization. Ninety-six hours following the 5th treatment of METH (P78), the rats received one dose of 0.5 mg/kg METH (s.c.) to reintroduce behavioral sensitization. Hyperlocomotion is a critical index caused by drug abuse, and METH administration has been shown to produce remarkable locomotor-enhancing effects. Therefore, a random forest model was used as the detector to extract the feature interaction patterns among the collected high-dimensional locomotor data. Our approaches identified neonatal systemic LPS exposure dose and METH-treated dates as features significantly associated with METH-induced behavioral sensitization, reinstated behavioral sensitization, and perinatal inflammation in this experimental model of drug addiction. Overall, the analysis suggests that the implementation of machine learning strategies is sensitive enough to detect interaction patterns in locomotor activity. Neonatal LPS exposure also enhanced METH-induced reduction of dopamine transporter expression and [ 3 H]dopamine uptake, reduced mitochondrial complex I activity, and elevated interleukin-1β and cyclooxygenase-2 concentrations in the P78 rat striatum. These results indicate that neonatal systemic LPS exposure produces a persistent dopaminergic lesion leading to a long-lasting change in the brain reward system as indicated by the enhanced METH-induced behavioral sensitization and reinstated behavioral sensitization later in life. These findings indicate that early-life brain inflammation may enhance susceptibility to drug addiction development later in life, which provides new insights for developing potential therapeutic treatments for drug addiction., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Critical-sized cranial bone defects fail to re-ossify and require the surgical intervention of cranioplasty. To achieve superior bone healing in such cases, a hydrogel consisting of an interpenetrating network of collagen and elastin-like polypeptide to encapsulate bone morphogenetic protein-2 (BMP-2), doxycycline, and 45S5 Bioglass is developed. This hydrogel has an appropriate elastic modulus of 39 ± 2.2 kPa to allow proper handling during implantation. The hydrogel promotes human adipose-derived stem attachment, proliferation, and differentiation toward the osteogenic lineage, including the deposition of hydroxyapatite particles embedded within a collagenous fibrillar structure after 21 days of in vitro culture. After eight weeks of implantation of the acellular hydrogel in a critical-sized rat cranial defect model, only a small quantity of various pro-inflammatory (< 20 pg mg -1 ) and anti-inflammatory (< 10 pg mg -1 ) factors in the adjacent cranial tissue is noticed, indicating the overall biocompatibility of the hydrogel. Scanning electron microscopy evidenced the presence of new fibrous extracellular matrix and mineral aggregates at the defect site, with calcium/phosphorus ratio of 0.5 and 2.0 by eight and twelve weeks, respectively. Microcomputed tomography (Micro-CT) and histological analyses showed formation of mature mineralized tissue that bridged with the surrounding bone. Taken together, the acellular composite hydrogel shows great promise for superior bone healing after cranioplasty., (© 2022 Wiley-VCH GmbH.)

Background Opioid sparing anesthesia and enhanced recovery after surgery protocols are not innovative ideas. However, the utilization of pancreaticoduodenectomy is limited. With the rise in awareness of the opioid epidemic in the United States, we have created a multimodal approach to anesthesia and postoperative care to limit adverse effects of opioids and curb the use of opioids postoperatively. Methods We conducted a retrospective cohort study performed by chart review of an opioid-sparing anesthetic and enhanced recovery after surgery (ERAS) protocol initiated jointly by the anesthesiology departments and transplant surgery for pancreaticoduodenectomy from January 2017 to October 2019. Results Demographic data was found to be comparable between the control and protocol groups. Hospital length of stay, ICU length of stay, and opioid requirements significantly decreased in the protocol group. Hospital length of stay decreased from 8.92 to 5.72 days, ICU days decreased from 1.52 to 0.42 days, and narcotics for the first five hospital days were significantly decreased from 130.13 to 71.2 morphine milligram equivalents. Conclusion Proper postoperative pain management can improve patient satisfaction and decrease complication rates. Pancreaticoduodenectomy is a complicated procedure with relatively limited data regarding enhanced recovery after surgery protocols. Likewise, there is limited data regarding opioid-sparing anesthesia techniques. Our protocol produced promising hospital length of stay and reduced opioid administration during the first five hospital days without increasing 30-day readmission rates., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2021, Leech et al.)

Background In the USA, spinal fusion surgery incurs the highest hospital cost. Despite the recent advances in the application of enhanced recovery after surgery (ERAS) protocols in these surgeries, the efficacy of these protocols in improving the perioperative outcomes remains unclear. We conducted a retrospective review as a quality improvement (QI) project to analyze the efficacy of the ERAS protocol with intraoperative modified thoracolumbar interfascial plane (mTLIP) block to determine whether these interventions reduce the length of stay (LOS) and opioid requirements during the postoperative period. Methods Retrospective reviews of adult patients (>18 yrs) who underwent elective lumbar spinal fusion or laminectomy at our institute were reviewed. Patients were administered oral gabapentin and acetaminophen preoperatively. Prior to incision, an mTLIP block was performed using liposomal bupivacaine. Intraoperatively, ketamine, ketorolac, and tranexamic acid were administered. Postoperative, pain control was treated with scheduled acetaminophen, ketorolac, and low-dose ketamine infusion. Hydromorphone and oxycodone were administered for breakthrough pain. Patients who underwent a similar procedure without ERAS protocol were chosen as controls to assess the efficacy of ERAS protocol. Data pertaining to patient demographics, operative and perioperative use of analgesics, LOS, 90-day readmissions, and morbidity were collected. Patients who underwent laminectomy and spinal fusion surgery were analyzed separately Results A total of 65 patients were identified; laminectomy (n- 24), spinal fusion surgery (n-41). In the laminectomy patients, treatment group (n-12) and the control group (n-12). Treatment group receiving the ERAS protocol with the regional anesthesia via the mTLIP (n= 12) opioid requirement was reduced by 51.42% [P = 0.03], and LOS was reduced by 2.04 days [P = 0.01] [0.75 days vs. 2.79 days]). In the spinal fusion patients, treatment group (n-15) and control group (n-26). Treatment group receiving the ERAS protocol with the use of regional anesthesia via the mTLIP group (n= 15), opioid requirement was reduced by 38.33% [P = 0.04]. No difference in LOS was observed at 5.4 days vs. 4.88 days (P = 0.28). Conclusion ERAS protocol in patients undergoing lumbar spinal surgery incorporated the use of regional anesthesia via the mTLIP block, we observed there is a statistically significant reduction in the LOS for lumbar laminectomy and a significant reduction in opioid administration for lumbar laminectomies and spinal fusion surgery., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2021, Kurnutala et al.)

Nearly all persons with spinal cord injury (SCI) will develop osteoporosis following injury, and further, up to 50% of all persons with SCI will sustain a fracture during their lives. The unique mechanisms driving osteoporosis following SCI remain unknown. The cannabinoid system modulation of bone metabolism through cannabinoid 1/2 (CB1/2) has been of increasing interest for the preservation of bone mass and density in models of osteoporosis. Using a thoracic vertebral level 8 (T8) complete transection in a mouse model, we performed daily treatment with a selective CB2 receptor agonist, HU308, compared with SCI-vehicle-treated and naïve control animals either immediately after injury for 40 days, or in a delayed paradigm, following 3 months after injury. The goal was to prevent or potentially reverse SCI-induced osteoporosis. In the acute phase, administration of the CB2 agonist was not able to preserve the rapid loss of cancellous bone. In the delayed-treatment paradigm, in cortical bone, HU308 increased cortical-area to total-area ratio and periosteal perimeter in the femur, and improved bone density in the distal femur and proximal tibia. Further, we report changes to the metaphyseal periosteum with increased presence of adipocyte and fat mass in the periosteum of SCI animals, which was not present in naïve animals. The layer of fat increased markedly in HU308-treated animals compared with SCI-vehicle-treated animals. Overall, these data show that CB2 agonism targets a number of cell types that can influence overall bone quality., Competing Interests: No conflicting financial interests exist., (© Michelle A. Tucci et al., 2021; Published by Mary Ann Liebert, Inc.)

Previous studies have demonstrated that pioglitazone, a peroxisome proliferator-activated receptor gamma (PPARγ) agonist, inhibits ischemia-induced brain injury. The present study was conducted to examine whether pioglitazone can reduce impairment of behavioral deficits mediated by inflammatory-induced brain white matter injury in neonatal rats. Intraperitoneal (i.p.) injection of lipopolysaccharide (LPS, 2 mg/kg) was administered to Sprague-Dawley rat pups on postnatal day 5 (P5), and i.p. administration of pioglitazone (20 mg/kg) or vehicle was performed 5 min after LPS injection. Sensorimotor behavioral tests were performed 24 h after LPS exposure, and changes in biochemistry of the brain was examined after these tests. The results show that systemic LPS exposure resulted in impaired sensorimotor behavioral performance, reduction of oligodendrocytes and mitochondrial activity, and increases in lipid peroxidation and brain inflammation, as indicated by the increment of interleukin-1β (IL-1β) levels and number of activated microglia in the neonatal rat brain. Pioglitazone treatment significantly improved LPS-induced neurobehavioral and physiological disturbances including the loss of body weight, hypothermia, righting reflex, wire-hanging maneuver, negative geotaxis, and hind-limb suspension in neonatal rats. The neuroprotective effect of pioglitazone against the loss of oligodendrocytes and mitochondrial activity was associated with attenuation of LPS-induced increment of thiobarbituric acid reactive substances (TBARS) content, IL-1β levels and number of activated microglia in neonatal rats. Our results show that pioglitazone prevents neurobehavioral disturbances induced by systemic LPS exposure in neonatal rats, and its neuroprotective effects are associated with its impact on microglial activation, IL-1β induction, lipid peroxidation, oligodendrocyte production and mitochondrial activity.

Study Objective: The aim of this study is to see if there are any clinical differences between using 35 F DLT for all patients versus using patient height regardless of gender to estimate appropriate DLT size., Design: Prospective randomized study., Setting: University Hospital., Patients: 50 patients age ≤18 years, undergoing lung or esophageal surgery requiring OLV., Interventions: Patients randomized to two groups (group-35F, group -DLT based on height)., Measurements and Main Results: Data collected include demographics, ASA status, airway assessment, number of intubation attempts, Cormack-Lehane grade, number of times DLT repositioned, incidence of sore throat, oxygen saturation at induction and oxygen saturation at 5 minutes and 10 minutes after OLV. There was no statistically significant difference in demographics, ASA classification, Mallampati score, number of intubation attempts, Cormack-Lehane grade, number of times DLT was repositioned, and incidence of sore throat. In height based DLT group the odds were higher for the incidence of sore throat in 37-41 F group. Oxygen saturation at induction, 5 minutes and 10 minutes after OLV are not statistically significant between the two groups. Conclusion: Our findings suggest that the majority of patients receive unnecessarily large DLTs for thoracic surgery, which not only makes intubation inherently more difficult but also increases their risk of postoperative sore throat.

Spinal cord injury (SCI) results in perturbations to the immune system leading to increased infection susceptibility. In parallel, the consumption of high-fat diets (HFD) leads to a chronic inflammation in circulation and body tissues. We investigated the impact of 16 weeks of HFD on chronically-injured rats. SCI rats under both chow and HFD showed peripheral leukocyte changes that include reduced percentages of total, helper and cytotoxic T, and natural killer cells. Expression of immune-related genes in the spleen and thymus reflected the impact of both chronic injury and diet. Changes to the immune system following SCI are adversely impacted by HFD consumption., Competing Interests: Declaration of Competing Interest Authors have no competing interests to declare., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Perinatal inflammation-induced reduction in pain threshold may alter pain sensitivity to hyperalgesia or allodynia which may persist into adulthood. In this study, we investigated the anti-inflammatory protective effect of interleukin-1 receptor antagonist (IL-1ra), an anti-inflammatory cytokine, on systemic lipopolysaccharide (LPS)-induced spinal cord inflammation and oxidative stress, thermal hyperalgesia, and mechanical allodynia in neonatal rats. Intraperitoneal (i.p.) injection of LPS (2 mg/kg) or sterile saline was performed in postnatal day 5 (P5) rat pups, and IL-1ra (100 mg/kg) or saline was administered (i.p.) 5 min after LPS injection. Pain reflex behavior, spinal cord inflammation and oxidative stress were examined 24 h after LPS administration. Systemic LPS exposure led to a reduction of tactile threshold in the von Frey filament tests (mechanical allodynia) and pain response latency in the tail-flick test (thermal hyperalgesia) of P6 neonatal rats. Spinal cord inflammation was indicated by the increased numbers of activated glial cells including microglia (Iba1+) and astrocytes (GFAP+), and elevated levels of pro-inflammatory cytokine interleukin-1β (IL-1β), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2) 24 h after LPS treatment. LPS treatment induced spinal oxidative stress as evidenced by the increase in thiobarbituric acid reactive substances (TBARS) content in the spinal cord. LPS exposure also led to a significant increase in oligodendrocyte lineage population (Olig2+) and mature oligodendrocyte cells (APC+) in the neonatal rat spinal cord. IL-1ra treatment significantly reduced LPS-induced effects including hyperalgesia, allodynia, the increased number of activated microglia, astrocytes and oligodendrocytes, and elevated levels of IL-1β, COX-2, PGE2, and lipid peroxidation (TBARS) in the neonatal rat spinal cord. These data suggest that IL-1ra provides a protective effect against the development of pain hypersensitivity, spinal cord inflammation and oxidative stress in the neonatal rats following LPS exposure, which may be associated with the blockade of LPS-induced pro-inflammatory cytokine IL-1β., (Copyright © 2020. Published by Elsevier Ltd.)

The goals of this study are to evaluate the ability of the multicomponent collagen-elastin-like polypeptide (ELP)-Bioglass scaffolds to support osteogenesis of rat mesenchymal stem cells (rMSCs), demonstrate in vivo biocompatibility by subcutaneous implantation in Sprague-Dawley rats, monitor degradation noninvasively, and finally assess the scaffold's ability in healing critical-sized cranial bone defects. The collagen-ELP-Bioglass scaffold supports the in vitro osteogenic differentiation of rMSCs over a 3 week culture period. The cellular (rMSC-containing) or acellular scaffolds implanted in the subcutaneous pockets of rats do not cause any local or systemic toxic effects or tumors. The real-time monitoring of the fluorescently labeled scaffolds by IVIS reveals that the scaffolds remain at the site of implantation for up to three weeks, during which they degrade gradually. Micro-CT analysis shows that the bilateral cranial critical-sized defects created in rats lead to greater bone regeneration when filled with cellular scaffolds. Bone mineral density and bone microarchitectural parameters are comparable among different scaffold groups, but the histological analysis reveals increased formation of high-quality mature bone in the cellular group, while the acellular group has immature bone and organized connective tissue. These results suggest that the rMSC-seeded collagen-ELP-Bioglass composite scaffolds can aid in better bone healing process., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background and Objective: No previous study has directly compared the levels of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) between smokers and individuals with diabetes mellitus (DM) with periodontitis. Therefore, the aim of this study was to evaluate the gene expression of MMP-1, MMP-2, MMP-8, MMP-9, TIMP-1 and TIMP-2 in tissues with chronic periodontitis (ChP) of smokers and individuals with type 2 DM., Material and Methods: Gingival biopsies were harvested from: non-smokers and non-diabetic individuals with ChP (n = 18) (ChP group); non-diabetic smokers (≥ 10 cigarettes per day for at least the past 5 years) with ChP (n = 18) (SChP group); non-smoking individuals with type 2 diabetes (glycated hemoglobin levels ≥ 7.5%) and ChP (n = 18) (DMChP group). The tissue levels of mRNA of MMP-1, MMP-2, MMP-8, MMP-9, TIMP-1 and TIMP-2 were evaluated by quantitative real-time polymerase chain reaction., Results: The MMP-8 expression was the lowest in the ChP group (p < 0.05). The DMChP group presented increased mRNA levels of MMP-2 and MMP-9, when compared to the SChP group (p < 0.05). MMP-1 expression and the MMP-1/TIMP-1, MMP-2/TIMP-1, MMP-8/TIMP-1, MMP-9/TIMP-1, MMP-1/TIMP-2 and MMP-2/TIMP-2 ratios were higher in the DMChP group than in the ChP and SChP groups (p < 0.05). The DMChP group presented lower mRNA levels of TIMP-1 than the ChP group (p < 0.05). The MMP-8/TIMP-2 ratio was the highest in the SChP group (p < 0.05)., Conclusion: Uncontrolled type 2 DM upregulates the ratio of MMP/TIMPs in sites with ChP more than smoking, which may contribute to a greater extracellular matrix degradation and periodontal breakdown in DM-related periodontitis., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Density variations, due to particle size and time and temperature of sintering, affect the delivery profile of substances from ceramic delivery devices. This investigation was conducted to study the effect of polylactic acid (PLA) impregnation on the porosity of hydroxyapatite (HA) capsules by studying the delivery rate of bovine serum albumin (BSA) and testosterone (TE). HA capsules were fabricated by cold compressing calcined particles at 1000, 3000, and 5000 lbs. Each group was subdivided into PLA-impregnated and non-PLA impregnated capsules. Each capsule was loaded with either 40 mg of TE (impregnated or nonimpregnated with PLA polymer) or BSA. Data obtained in this study suggest that: (1) PLA impregnation of HA ceramic capsules decreases the rate of release of drugs from the ceramic reservoir, (2) Physiochemical characteristics of the drugs to be delivered is an instrumental key in the sustained release profiles, (3) Compression load of HA reservoirs is a key factor in predicting the duration and level of sustained delivery, and (4) Polymer coating of HA ceramic capsules reduces the pore size, as well as, blocking some of the pores on the surface.

Demineralized bone matrix protein (DBM) was considered highly effective in stimulating bone healing. The objective of the study was to explore the use of tricalcium phosphate (TCP) delivery system to continuously deliver DBM in an osteoporotic condition and to evaluate changes in bone density and preservation of the spine. Ovariectomized Sprague Dawley rats were divided into three equal groups (n=16 per group). Animals in group I served as control, animals in groups II and III were surgically implanted with either empty (SHAM) or DBM filled TCP implants adjacent to L4/L5. Eight animals from each group were euthanized at 2 and 8 weeks post implantation. Femurs were evaluated for changes in density, and the lumbar spine was evaluated for changes in the endplate. Results of this study revealed (1) TCP implants were capable of delivering DBM for long duration, (2) use of sustained delivery of DBM did not induce untoward effects in the vital organs or in the uterus, fallopian tubes, or vaginal tissues, (3) DBM had no effect on chondrocyte differentiation in the spine, and (4) DBM did not increase bone density in osteoporotic female rats.

The needs of dental patients are becoming ever complex. This is largely due to the increasing number of systemic conditions that patients present with during routine or emergency dental visits, such as hypertension and angina pectoris. Medications that patients are prescribed have potential to cause effects on the oral tissues. There is a need to understand the interactions between the oral tissues and materials used to restore teeth. The objective of this study was to assess the physiological function of human gingival fibroblasts upon exposure to dental adhesives; polymethyl methacryate, OptiBond®, and Prime & Bond®; Porphyromonas gingivalis lipopolysaccharide and Cortisol or Nifedipine. An increase in metabolic activity was noted with the experimental group exposed to the combination of Prime & Bond®, LPS-PG, and Nifedipine (P<0.001). Oxidative stress was observed in experimental groups exposed to the combinations of PMMA, LPS-PG, and Cortisol (P<0.001); also noted in all three dental adhesives in combinations with LPS-PG and Nifedipine (P<0.001). Cellular membrane damage was evident in experimental groups exposed to the combinations of PMMA, LPS-PG, and Cortisol; PMMA, LPS-PG, and Nifedipine; and Prime & Bond®, LPS-PG, and Nifedipine. In conclusion, it appears that the addition of Nifedipine in combination with LPS-PG negatively alters cellular function by causing membrane damage and inducing oxidative stress.

The objective of this study was to investigate the synergistic effect of Thymoquinone (TQ) and Epigallocatechin-3-gallate (EGCG) using Caov-3 cell line as a model. A total of 144 wells were plated with 10^5 Caov-3 ovarian cancer cells. The wells were divided into groups of 72 wells for conventional and sustained delivery, respectively. Each group was subdivided into 4 groups of 6 wells. Group 1 served as control and groups 2, 3, and 4 were treated with TQ (16 µM), EGCG (3 µg/ml), and TQ plus EGCG, respectively. Biomarkers evaluations were performed following standard lab techniques. The results of the study revealed: conventional delivery of TQ, EGCG, and the combination did not induce intracellular oxidative stress (glutathione levels) at 24, 48, and 72 hours; however, sustained delivery of all agents showed significant difference from the control at 72 hours; and there was no significant difference at 24 and 48 hours. Overall conclusion: the route of potent agents to manipulate ovarian cancer cells is highly dependent on the route of administration.

The FaDu squamous cell carcinoma cell line was used to test combination treatments of natural (EgCg and TQ) drugs and a chemotherapeutic (5-FU) drug to determine if combining drugs with different cell cycle targets would be more effective at destroying cancer cells than one drug that only targets one phase of the cell cycle. There were a total of four different combinations. One combination consisted of all three treatments: EgCg, TQ, and 5-FU. The other three combinations were as follows: EgCg + TQ, EgCg + 5-FU, and TQ + 5-FU. The combination treatments were measured at 24, 48, and 72 hours. There were significant reductions in cell number at each time increment. The combination of any two drugs especially a natural drug plus a chemotherapeutic drug proved to be more effective than single individual drugs in eradicating cancer. The combination of EGCG + TQ + 5-FU proved to be most effective in cell reduction. Since a combination of drugs target more than one phase of the cell cycle, the cells were less likely to become resistant to two or more drugs.

Surgically repaired tendons are plagued by complications related to the healing response. Adhesion formation between the tendon and its sheath or surrounding tissues inhibits free gliding and results in a loss of excursion. The random orientation of collagen deposition at the site of repair creates a focal area of weakness, rendering the tendon prone to rupture at this point. Even the strongest, most technically precise repair can be negated by either of these complications. Research in tendon injury and healing is primarily focused on the pathogenesis of fibrosis and how to prevent the formation of excessive scar tissue. However, no widely accepted therapy currently exists to promote healing and prevent fibrosis in surgically repaired tendons. The goal of this study was to determine the effects of varying concentrations of transforming growth factor ß1(TGF-ß1), the main constituent of a clinically used treatment, PRP, on fibroblast cells. Cells were treated with low, medium, and high concentrations of TGF-ß1 for periods of 24, 48, and 72 hours and cell proliferation, damage, and morphology were evaluated at each time point. The results show that low dose treatment resulted in significant increases in cell number and cell along with distinct cytological changes within 48 hours of treatment. This information is clinically important and may help to provide an explanation for the inconsistencies seen with use of PRP as an adjuvant for tendon and ligament healing.

Osteoporosis and cardiovascular disease (CVD) are common age-related conditions, which are major public health problems leading to an increase in mortality, morbidity, and disability. There have been several connections found between CVD and osteoporosis such as common genetic factors, risk factors, and pathological mechanisms. There is a direct effect of estrogen on CVD and osteoporosis that is demonstrated by the manifestation of estrogen receptors on osteoblasts, osteoclasts, and vascular endothelial and smooth muscle cells. Loss of estrogen has been found to be involved in the pathogenesis of atherosclerosis and bone loss through modulation of other factors including cytokines and oxidized lipids. The goal of this proposed research was to determine if sustained delivery of estrogen is capable of regulating bone cell function while improving cardiovascular panels. Ovariectomized Sprague Dawley rats were administered estradiol at a rate of 5ng/day over an eight-week period. Body weights, estradiol levels, cholesterol levels, and bone strength were determined at 2, 4, and 8 weeks following sustained delivery of estradiol and compared with intact control and ovariectomized control animals. Estrogen replacement resulted in improved cholesterol panels without significant changes in bone flexural strength or improvements in bone porosity. Additional long term studies are needed to determine if the benefits of estrogen replacement outweigh the inherent risks associated with hormone replacement therapies.

The need for bone graft substitutes is increasing. The understanding of how these potential treatments exert their effect is lacking. Currently, BMP’s are used clinically for augmentation of fracture healing and the exact mechanism(s) how they contribute bone healing is not clear. The literature show that demineralized bone matrix protein, OP-1, and IGF-1 are all factors which contribute to increased bone growth. The goal of this study was to determine if these factor signal through similar pathways that can affect bone growth. Our findings show upregulated intracellular MAPK and JAK STAT signaling by OP-1 as well as novel upregulated intracellular SMAD signaling by IGF-1. As expected, OP-1 signaled through the SMAD pathway and IGF-1 signaled through the MAPK and JAK STAT pathways. DBM which contains both OP-1 and IGF-1 signaled through all three pathways as well. The exact role of each of these factors needs to be further investigated to determine which pathway is crucial for normal bone formation and which pathways will be important for bone remodeling.

This study investigated the effects of dual delivery of statin and vancomycin on angiogenesis during the healing process of a femoral defect injury using tricalcium phosphate lysine (TCPL) delivery system in an animal model. The experimental design consisted of 14 rats divided into the following three groups: Group I animals (n=5) served as the intact control without treatment. Group II animals (n=5) were subjected to a surgically induced defect (2 mm, midshaft of the right femur) and implanted (IM) with TCPL capsules loaded with vancomycin (20mg) (TCPL-AB). Group III animals (n=4) were operated on in a similar fashion as Group II, and subsequently implanted with TCPL capsules loaded vancomycin (20 mg) plus statin (5 mg). The animals were euthanized at 30 days post-implantation using overdose of isoflourane. The right femurs were then harvested in addition to the vital organs, the reproductive organs, and sample of the adjacent skeletal muscles. The hard and soft tissues were evaluated histopathologically by following laboratory standard techniques. The results of this study indicated that statin plus vancomycin treated animals had increased angiogenetic activities with many blood vessels compared to the sham group and the animals also healed in a greater magnitude than the sham group (independent evaluators (p<0.001)). Histomorphometric analysis demonstrated that exposure to sustained delivery of statin resulted in increased blood vessels. It appeared there is a direct correlation of the increased angiogenesis and the increased bone formation in the statin group and this may be one of the mechanisms with which statin form bone. In conclusion, data obtained from this study demonstrated that sustained delivery of statin by TCPL resulted in a remarkable increase in angiogenic and osteogenic activities during the healing process of a femoral defect.

Dental adhesives are necessary for the retention of specific dental restorations utilized to repair the anatomy of the tooth after dental decay is removed. Adhesives come into contact with healthy and diseased periodontal tissues. Porphyromonas gingivalis is a gram negative bacterial pathogen, and lipopolysaccharide (LPS-PG) is an endotoxin found in gingival connective tissues of patients who suffer from periodontal disease. The presence of the endotoxin causes inflammation. This study aims to evaluate the effectiveness of potent dental adhesives when human gingival fibroblasts are challenged with LPS-PG. The fibroblasts were exposed to the dental adhesives polymethly methacrylate (PMMA), OptiBond®, and Prime & Bond® which were purchased from Patterson Dental, a national dental materials supplier. The human gingival fibroblasts (HGF-1, ATCC® CRL-2014™) were purchased from American Type Culture Collection (ATCC). The porphyromonas gingival lipopolysaccharide (LPS-PG) was purchased from Fisher Scientific (Pittsburg, PA). No significant differences in metabolic behavior was detected among the groups (p<0.132). While the glutathione assay determined that there was not any significant increase in oxidative stress levels; the lactate dehydrogenase assay identified significant cellular damage in the group exposed to combinations of the Prime & Bond® adhesives and LPS-PG at 48 hour intervals (p<0.003). No significant changes were noted in cellular morphology at any phases, and all cells demonstrated typical fibroblast spindle shape.

Approximately 7.9 million bone fractures occur every year in the United States, with an estimated cost of $70 billion. It is estimated that 5-10 percent of patients will develop infections or delayed healing following orthopedic surgery. The probability of osteomyelitis is related to the severity of trauma and infection is most frequent after high velocity, open injuries with skin loss. Complications with bone healing and antibiotic drug-resistance increases morbidity and costs associated with osteomyelitis are estimated at $15,000-$30,000 per incident. Traditional osteomyelitis treatment often requires additional surgery and systemic antibiotics that are harmful, burdensome, and costly. There is limited evidence in the literature examining the effects pharmacological agents tobramycin (TOB) and parathyroid hormone (PTH) have on the mesenchymal stem cells (MSCs) that differentiate into many cell types. The purpose of this investigation was to evaluate the in vitro dose-dependent effects of TOB and PTH on mesenchymal stem cells. MSCs were treated for periods of 24, 48 and 72 hours in the presence of TOB and PTH. Following incubation, MSCs were examined for cellular morphology, viability, proliferation, and damage. Our results show an increase in morphological changes following treatments with both PTH and TOB which are confirmed by decreases in cellular protein concentration. Are results also indicate PTH acts as an essential factor in mesenchymal development and may control differentiation into osteogenic or chondrogenic lineage. Development of targeted delivery of PTH and TOB to a fracture may be effective at treating both infection and bone loss.

Ovarian cancer is the fifth most common cancer and the leading cause of mortality among the gynecologic cancers. Ovarian cancer is a very devastating disease it is rarely diagnosed in its early stages, and it is usually quite advanced by the time diagnosis is made leading to very poor outcomes. It is believed that in advanced stage ovarian cancer cells produce a protein that enables the tumor cells to proliferate and become resistance to conventional drug treatments. Development of new treatment options strongly relies on understanding the ovarian carcinoma ability to proliferate and to attack the cells using combination drug treatments that are synergistic. Recent studies in our laboratory have indicated that the exposure of Thymoquinone (TQ) and Epigallocatechin-3-gallate (EGCG) to adenocarcinoma of the prostate, colon, and pancreas. This study was executed to investigate the effectiveness of TQ and EGCG on reducing early stage ovarian cancer cells. A total of 72 wells were plated with (105) Caov-3 ovarian cancer cells according to standard lab protocols. Each group was subdivided into 4 groups. Group 1 served as control and groups 2, 3, and 4 were treated with TQ (16 µM), EGCG (3 µM), and TQ + EGCG, respectively. Biomarkers and morphological evaluations were performed following standard lab techniques. The results of the study revealed: (1) an increase in nitric oxide following administration of EGCG and the combination at 24 and 48 hours (p<0.05) and (2) no membrane or cellular damage to the cells at all phases. The results of this study suggest used in combination have shown to be more effective method for manipulating Caov-3 ovarian cancer cells than when used alone.

Patients with type 2 diabetes mellitus require adequate blood glucose control with insulin, metformin or other anti-hyperglycemic agents to prevent significant co-morbidities such as cardiovascular disease. Studies have determined that metformin increases the activity of AMP activated protein kinase and reduces hepatic glucose production and increases tissue sensitivity to insulin. Insulin has some vascular protective qualities; however insulin resistance of type 2 diabetes may be harmful to vascular endothelium. The objective of this study was to investigate the effects of metformin and insulin on H9c2 rat cardiomyocytes in high glucose culture media. Results of this study indicate that clinical doses both insulin and metformin are capable of reducing supernatant glucose levels in high glucose media. Metformin may offer some protection to cardiomyoctyes. Morphologically, cellular changes occurred after exposure to both insulin and metformin at both 24 and 48 hours although at 72 hours, both groups exhibit evidence of cellular damage.

The experimental impact of retinoic acid (All Trans Retinoic Acid; ATRA), citrals, ovalbumin and mold spores in the development of lung pathology and tissue remodeling are not well established in the literature. As well, the role of these agents in lung pathology was not ascertained under an in vivo setting. Therefore, it is hypothesized that citrals, ATRA, ovalbumin and mold-spore exposure will sensitize lung tissues and will lead to the development of lung tissue pathology in these animals. The study used an IACUC approved between-subject in vivo randomized split plot factorial design (F344 rat model; N=30). Mold spores were applied to animals by intra-tracheal route whereas vehicle, ovalbumin, C1, C2 and ATRA were administered by intra-peritoneal route. Rat weight data and blood were collected on Days 1 and 21. All animals were sacrificed on day 21 and lung tissues were processed for histopathology. Evidence from weights and blood (ANOVA and Duncan) as well as histopatholgical analysis supported the findings that exposure of these animals to C1, C2, ATRA, ovalbumin and mold spores showed different levels of lung tissue damage representing environmental exposure to these agents. This promising study showed variable lung tissue responses to the administration of ATRA, ovalbumin, citrals, and mold spores in the development of various levels of lung tissue pathologies.

The impact of retinoic acid (All Trans Retinoic Acid; ATRA) in the development of lung pathology and tissue remodeling are not well established in the literature. As well, the role of citral (inhibitor of retinoid function) in the improvement of lung pathology was not ascertained under an in vivo setting. Therefore, it is hypothesized that ATRA and ovalbumin exposure will sensitize lung tissues leading to lung tissue pathology and that citrals (C1 and C2) will reverse or ameliorate the related pathological damage to lung tissues. The study used an IACUC approved between-subject in vivo randomized split plot factorial design (F344 rat model; N=40). Animals were exposed to 8 different treatments including vehicle, OVA, ATRA, citrals (C1 and C2) and their ovalbumin combinations (OVA+ ATRA, OVA+ C1, and OVA+ C2) by intra-peritoneal route. Rat weight data and blood were collected on Days 1 and 21, all animals were sacrificed on day 21 and lung tissues were processed for histopathology. Results from weights and blood (ANOVA and Duncan) as well as from the histopatholgical analysis supported the findings that exposure of F344 rats to OVA combinations with ATRA and citrals showed various levels of lung tissue damage that was improved or worsened by either C1 or C2. This promising study showed variable responses on the interaction of ovalbumin, citrals, and ATRA as related to their damage/improvement of related lung tissue pathologies.

The objective of this investigation was to evaluate the location and activity of cytokines in the fibrous tissue surrounding tricalcium phosphate (TCP) implants loaded with androgenic hormones. Sixteen animals in four experimental groups (n = 4/group) were implanted with one TCP implant each: Group I (control), Group II (testosterone), Group III (dihydrotestosterone), and Group IV (androstenedione). At 90 days post-implantation, the fibrous tissue surrounding the implants were evaluated following staining with antibodies to IL-1ß, IL-2, IL-6, and TNF?. Data were collected on the presence and distribution of cytokines within the fibrous tissue surrounding all four groups. IL-1ß was primarily found intercellular and associated with fibroblasts and macrophages of Groups I-III. IL-2 was present in the extracellular matrix and was sporadically found on the surface of macrophages in Groups I-III. IL-6 was found primarily concentrated in the fibroblast and collagen rich portions of the fibrous tissue matrix in Groups I-III. TNF-? was present in the extracellular matrix of the fibrous tissue of all four groups and was strongly associated with fibroblast and macrophage rich areas. The results of this study confirm activity of cytokines on target cells and indicate their actions may vary in their effect within the fibrous tissue surrounding TCP implants loaded with androgens.

Alzheimer’s disease (AD) is the only cause of death among the top 10 diseases that cannot be prevented, cured, or slowed with the current treatments available. Amyloid beta is thought to be the main initiator of AD cognitive decline, activating internal pathways which lead to inflammation, oxidation, and cell death. The objective of this study was to determine if pretreatment of human SH-SY5Y neuroblastoma cells, a model for AD, with antioxidants thymoquinone (TQ), epigallocatechin-3-gallate (EGCG), or dilinoleoylphosphatidylcholine (DLPC) 30 minutes prior to a challenge with tumor necrosis factor a (TNFa), an inflammatory mediator, can prevent oxidation of amyloid beta (Aß). Following treatment, cells were incubated and groups evaluated at 24, 48, and 72 hours. Human amyloid precursor protein (APP) enzyme-linked immune-sorbent assay (ELISA) and nitric oxide assays were performed from supernatant whereas protein and glutathione assays were performed from cells. When TNFa was added to cells, Aß significantly increased 3-fold compared to untreated cells. The addition of antioxidants EGCG, TQ, and DLPC reduced Aß back toward control value at the initial time point. TNFa also caused a significant increase in nitric oxide without changes in glutathione. TQ administered to the cells prior to a challenge with TNFa resulted in a decrease in nitric oxide and an increase in glutathione which may be a possible mechanism to reduce inflammation and reduce oxidation. Additional studies are needed to determine the signaling pathways implemented in the SH-SY5Y cells following TNFa.

Acute Lymphoblastic Leukemia remains the most common cancer for children, and if left untreated is rapidly fatal. The gold standard for treatment of ALL in children is with a class of drugs known as the antracyclines. Long term outcomes following treatment of leukemia with antracylines can result in cardiac abnormalities including arrhythmias, congestive heart failure, myocardial infarction, hypertension and left ventricular failure. Thymoquinone is a natural product that has demonstrated anti-proliferative, anti-inflammatory, anti-cancer, and chemo-protective effects in control trials as well as a reduction in cardiotoxicity in antracyline treated rats. The aims of the study were to determine if thymoqunione could be used to reduce leukemia cell viability without injuring primary cardiomyocyte, and to determine its effects if used in conjunction with a known chemotherapeutic agent. Cellular viability and morphological changes were observed in the, RAW leukemia cells and cardiac myocytes following treatment with thymoquinone, antracyline (doxorubicin), alone and in combination for 24, 48 and 72 hours. The results suggest that thymoquinone treatment in RAW leukemia cells reduced the cell number without altering the morphology, while doxorubicin reduced cell number and induced spindle cell formation and increased cellular damage. Findings also suggest RAW cell apoptosis increased in combination therapy with thymoquinone and doxorubicin. Thymoquinone administered to cardiomyocytes showed similar morphological changes as control over time in culture; whereas doxorubicin treated cells showed evidence of loss of connectivity and disruption of cell membranes. Combination treatment with doxorubicin and thymoquinone demonstrated significant cardiac myocyte survival at concentrations when used alone were able to reduce the leukemia cells. Overall, the data is promising and may provide a treatment regime to protect the heart tissue. Additional work is warrant to understand the mechanisms involved to reduce cardio toxicity by combining thymoquinone with doxorubicin.

Platelet dense granules contain serotonin, adenosine triphosphate (ATP), and adenosine diphosphate (ADP). These molecules are present in platelet rich plasma (PRP), and may therefore have an impact on the efficacy of PRP therapy. Additionally, nucleotides are important extracellular signaling molecules in a variety of tissue types including bone. The purpose of this investigation was to evaluate the in vitro dose-dependent effects of extracellular adenosine triphosphate (ATP) exposure on activity of human osteoblast-like cells. MG-63 cells were exposed to phosphate buffered saline (control group) or ATP solution (20 µM, 100µM, 200 µM). Osteoblast viability was evaluated at 24, 48, and 72 hours using nonspecific and osteoblast-specific markers and cellular morphology. No significant differences in total protein, malonlydialdehyde (MDA), or glutathione were observed with ATP exposure at any timepoint. High dose ATP exposure resulted in a significantly higher production of nitric oxide compared to controls and other groups. With respect to alkaline phosphatase activity and osteopontin production, no significant differences were present with ATP exposure. Overall conclusion: Extracellular ATP exposure modulated osteoblast activity with no change in cell viability in vitro.

The present study evaluated the morphology of human gingival fibroblasts when exposed to dental adhesives in combination with glucocorticoids or dihydropridines in an in vitro environment. The cultured gingival fibroblasts used were obtained from the American Type Culture Collection (ATCC). The three dental adhesives used were Polymethyl methacrylate, OptiBond®, and Prime & Bond®. The glucocorticoid of choice was Cortisol along with the dihydropridine (Nifedipine). Fibroblasts were treated with each bonding agent, Cortisol, or Nifedipine for durations of 24, 48, and 72 hours. Morphological evaluation revealed that at 24 and 48 hour phases, there were no observable differences between the control and groups exposed to a combination of an adhesive and Cortisol. However, at 72 hours the cells exposed to combinations of OptiBond® with Cortisol and Prime & Bond® with Cortisol displayed predominantly spindle shaped cells compared to the predominantly round cells observed in the other samples. Cells exposed to a combination of adhesives with Nifedipine at 24 hour phase did not display any significant morphological alterations. In contrast, cells exposed to Prime & Bond® in combination with Nifedipine at the 48 hour phase revealed occasional morphological changes compared to control group. Likewise and at the end of 72 hour phase, the OptiBond®/Nifedipine and Prime & Bond®/Nifedipine groups displayed morphological differences such as clustering compared to the control. Biochemical analysis determined that there was a significant difference among the groups exposed to adhesives with Cortisol at 48 (P<0.002) and 72 hours (P <0.001). Glutathione (GSH) and Malondialdehyde levels (MDA) indicated that there were significant differences in the metabolic activities in groups exposed to combinations of dental adhesives and Cortisol at all phases. There were no differences in lactate dehydrogenase (LDH) levels among the groups exposed to combinations of adhesives and Cortisol or adhesives and Nifedipine. It is concluded that there were minor structural (morphological) alterations and major functional (biomarkers) changes observed upon exposing the gingival fibroblasts in combinations with Cortisol (stress environment) or Nifedipine.

Platelets are naturally active in the healing process. Derived from megakaryocytes, platelets are not actual cells because they have no nucleus or DNA and cannot synthesize their own proteins. When platelets are activated by stress, contact with tissues, or thrombin and calcium they release many powerful factors through a process referred to as degranulation. Degranulation describes the release of factors contained within two storage areas of the platelet, alpha and dense granules. A few of the known components released from the alpha granules of activated platelets include PDGF, VEGF, EGF, FGF and TGF-ß. These factors perform various functions that contribute to inflammation, tissue healing and tissue regeneration. The dense granules release their contents prior to alpha granule release at the site of activation. Well classified dense granule constituents are ionized calcium, adenosine di-phosphate, adenosine tri-phosphate , histamine, epinephrine and serotonin. Most applications of platelet rich plasma (PRP) center around a theory that growth factors released from the alpha granules of platelets play a critical role in tissue healing following injury. There is limited evidence in the literature regarding the role of the dense granule components role during inflammation. Our goal is to define serotonin release from the dense granule and the response of activated macrophage to serotonin. Serotonin has very little effect on macrophage cells in terms of both viability and function. Serotonin administered to macrophage cells does not result in a reduction of cell number, inflammatory cytokine production, or induction of a nitric oxide burst on its own. In the presence of lipopolysaccharide (LPS) activated macrophages serotonin reduces the initial LPS induced nitric oxide burst and suppress LPS induced cytokine (IL-1) production by 72 hours. Overall, the data suggest the possibility that the dense granule factors may contribute to the initial stages of healing.

The incorporation of platelet-rich plasma (PRP) into scaffolds for application in musculoskeletal injuries has been a topic of recent interest in orthopaedic surgery. Platelets have dense granules containing ADP, ATP, serotonin, and calcium; and alpha granules containing PDGF, VEGF, IGF, TGF-ß, and EGF. Particular focus of previous studies has been on mitogenic effects of alpha granules, but the role of dense granules in PRP therapy currently remains undefined. The objective of the present study was to evaluate the effect of ATP, ADP, and serotonin upon osteoblast viability in vitro. Human osteoblast-like cells (MG-63 cells) were exposed to phosphate buffered saline (control group), ATP (20µM), ADP (10µM), and serotonin (11.75nM) for 24, 48, and 72 hours. Osteoblast viability was evaluated at each timepoint using biochemical assays. When compared to controls, osteoblasts treated with ATP and ADP resulted in a significant reduction in cell number, while serotonin caused an increase at 24 hours. Similar trends were noted at later timepoints. At 48 hours, a trend towards increase in glutathione was observed with ADP and ATP, but was not sustained at 72 hours. No significant differences in membrane damage were detected between groups. At 24 and 48 hours, ADP significantly increased nitric oxide production. Results of this study demonstrate that ATP, ADP, and serotonin induced significant structural adaptive responses to osteoblastic activities. The data revealed minimal functional alteration as evident by biomarker measurements. Overall conclusion: the results provided further insight regarding PRP therapy for traumatized bone.

In osteoarthritis, the metabolic activity of the chondrocytes is shifted toward a state where new matrix synthesis is outweighed by breakdown of matrix constituents. The result is degeneration and gradual loss of articular cartilage. Although osteoarthritis is frequently regarded as a non-inflammatory form of arthritis, considerable data implicates a role for pro-inflammatory cytokines in the cartilage destruction associated with osteoarthritis. The best studied pro-inflammatory cytokines in osteoarthritis are interleukin-1ß (IL-1ß), tumor necrosis factor-a (TNF-a), and interleukin-6 (IL-6). Since articular cartilage is not vascularized, it must rely on diffusion from the articular surface for nutrient and metabolic exchange. Consequently, the entire metabolism of the cell is geared towards operating at a low oxygen tension. In this study, chondrocytes were challenged with pro-inflammatory cytokines at 21% O2 and 6% O2. Chondrocyte proliferation, membrane integrity, oxidative stress, matrix metalloproteinase – 9 (MMP-9) and hypoxia inducible factor-1a (HIF-1a) production were measured. Our results showed that there was less of a decrease in cell number at 6% O2 compared to 21% O2 after they were challenged with pro-inflammatory cytokines. In addition, there was less of an increase in oxidative stress, membrane damage and MMP-9 production at 6% O2 compared to21% O2. The significance of this study represents the first attempt to replicate a diseased inflammatory environment characterized by an osteoarthritic joint in vitro and to examine these effects on the growth and stability of chondrocytes.

Hydrogen peroxide kills bacteria; however, control of a specific dose limited control over cellular response and function. In this study, the GOX/CAT system was used to independently provide and control the amount of H2O2 and oxygen in cell culture. The GOX/CAT system is composed of glucose oxidase (GOX) and catalase (CAT). Lipopolysaccharide (LPS) is molecules on the outer membrane of gram-negative bacteria. LPS elicits an inflammatory stimulus which can activate macrophages to produce pro-inflammatory cytokines, TNF-a, and specific mediators of inflammation including NO and prostaglandins. Macrophages are known to provide a prominent defense against bacteria. The aim of this experiment was to determine if GOX/CAT (10µMH2O2) plus conditioned media from LPS challenged macrophage cells will alter fibroblast cell numbers, viability and morphology after 24, 48, 78 hours. Human gingival fibroblast (HGF) CRL-2014 were treated with conditioned media from RAW 264.7 cells with 4µg/mL LPS, GOX/CAT (10µMH2O2) and 4µg/mL LPS + GOX/CAT (10µMH2O2) at 24, 48 and 72 hours. Cell protein, MDA and GSH levels and cell morphology were determined after each incubation periods. The results showed no significant difference in protein levels and MDA levels throughout the experiment. Reduced glutathione concentration in treated cells was statistically lower than control suggesting the possibility of intracellular damage. Overall, cell response to H2O2 did not alter cell proliferation and viability of LPS conditioned media treated fibroblast. Morphological results indicated a trend in possible adaptive response to LPS after 48 and 72 hours. Additional experiments are needed to determine chronic exposure on HGF.

The purpose of this study was to test the effectiveness of three potential bonding materials on the proliferation, viability, and functional capacity of the human gingival fibroblasts in an in vitro environment. The cultured gingival fibroblasts used were obtained from the American Type Culture Collection (ATCC). The three bonding materials used were polymethyl methacrylate (PMMA), OptiBond®, and Prime & Bond®. Fibroblasts were exposed to each bonding agent for durations of 24, 48, and 72 hours. Cellular protein levels determined that exposure to Prime & Bond® resulted in a significant increase in cellular protein (17.62±2.61 mg/mL) after 24 hours when compared to control and PMMA and OptiBond® exposures. Cellular protein levels were similar for experimental and control groups after 72 hours. The MDA assay showed no statistically significant differences at 24, 48, and 72 hours following exposure with PMMA or OptiBond® when compared to control cells. Membrane lipid peroxidation was not detected in the Prime & Bond® exposed cells. There is no significant decrease in cellular damage as measured by activity of LDH in the cell media at 24 and 48 hours. Prime & Bond® incubated with fibroblast cells for 72 hours resulted in a significant increase in LDH activity in the media (40.512 ± 4.437 IU/mg Protein) compared to control (19.975 ± 3.463 IU/mg Protein) and other dental bonding compounds. OptiBond® resulted in a decrease in reduced glutathione levels after 24 hours when compared to PMMA and Prime & Bond® exposure. No differences in intracellular levels were detected after 48 and 72 hours. This investigation suggests that the physiochemical characteristics of restorative dental adhesives have to be considered a key factor in the biocompatibility status.

Study Design:  Randomized controlled animal trial., Objectives:  To analyze the early and late phase effects of platelet-rich plasma (PRP) injection into and around the damaged intervertebral disc using an animal model, and to evaluate the needle puncture technique in creating a degenerative disc model., Methods:  The L4-L5 intervertebral disc of 18 adult Sprague-Dawley rats was injured with a 21-gauge needle. Animals received an immediate injection of PRP, or a delayed injection of PRP 2 weeks postinjury, or no further intervention (sham) (n = 6/group). Six uninjured controls were used. Magnetic resonance imaging (MRI) was performed for a control at time 0 and each group at 4 weeks postinjury, this allowed resolution of inflammation and an accurate assessment of healing. Specimens were collected from immediate PRP group at 2 and 4 weeks postinjury, and the delayed PRP group at 4 and 6 weeks postinjury. The sham and control specimens were collected at 2, 4, and 6 weeks postinjury. Each disc was sectioned and analyzed subjectively for overall structure, cellularity, and water content (utilizing MRI). Disc height was measured using descriptive statistics and one-way analysis of variance (ANOVA) with Tukey post hoc analysis. It was presumed both groups injected with PRP would have better outcomes than the sham group., Results:  The sham group had clear degenerative changes with loss of organizational structure, empty space, fibrous tissue, and inflammatory cells, indicating needle puncture leads to degeneration. The PRP treated groups had fibers that were damaged with empty spaces and inflammatory cells. However, there was maintenance of the ring structure and the nucleus appeared to have a healthy central portion. Overall, both PRP - treated groups retained more normal morphologic features, contained fewer inflammatory cells, and had higher fluid content on MRI; however the effect was more pronounced in the immediate injection group. The disc height was significantly different in the sham and immediate injection group at the 4-week interval., Conclusions:  This percutaneous needle puncture technique is an effective method for creating a degenerative disc model without posterior destabilization of the spine. The administration of PRP has a protective effect on damaged discs in the acute and delayed injection settings representing clinical treatment with PRP in the early versus late stages of the degenerative process. It appears that earlier intervention in the disease process would be more beneficial than PRP treatment of already severely degenerated discs.

Unlabelled: Hypoxia is the initial cellular insult in carpal tunnel syndrome. While much is known about the gross and histological progression of this prevalent disease, little is currently known about its molecular basis. Additionally, some debate presently exists regarding whether the disease occurs as the result of an inflammatory or a fibrotic pathology., Objective: The objective of these investigations was to determine the transcriptional and translational regulation in MRHF fibroblasts subjected to an hypoxic culture environment with respect to HIF-1a and TGF-ss1 mRNA and protein expression., Methods: MRHF fibroblasts were cultured under anoxic conditions for various durations up to 72 hours alongside normoxic controls. HIF-1a and TGF-ss1 mRNA levels relative to baseline were determined by real-time PCR. GAP-DH was used as the reference standard for normalization and relative quantification of fold-differences in mRNA expression was determined by the comparative (2[delta][delta]Ct) CT method. Relative differences in protein levels were determined by Western blotting for both HIF-1a and TGF- ss1., Results: Transcriptional and translational up-regulation of both HIF-1a and TGF-ss1 were observed in response to hypoxic culture conditions. Peak mRNA expression was observed after 15 min of anoxic exposure for both HIF-1a (7.3-fold increase) and TGF-ss1 (3.3-fold increase). In contrast, peak protein levels were observed much later in the course of hypoxia with HIF-1a and TGF-ss1 both demonstrating a significant increase over baseline levels after 2 hours., Conclusion: Both HIF-1a and TGF-ss1 have been shown to regulate the expression of connective tissue growth factor (CTGF). The pronounced and sustained up-regulation of HIF-1a suggests that CTGF expression in hypoxic carpal tissues is predominantly under the early regulatory influence of HIF-1a.

Study Design: A novel degenerative disc disease model and sustained delivery method for corticosteroid in male Sprague-Dawley albino rats., Objectives: To develop a model of degenerative disc disease and to determine the effect of continuous sustained release of corticosteroid on the process of degeneration within the traumatized disc., Summary of Background Data: The current modalities of treating symptomatic degenerative disc disease are either conservative or surgical. However, there is no cure for the degenerative process and prevention, therefore, is the ideal treatment. An understanding of the mechanisms involved in disc degeneration is crucial to develop new methods for prevention and treatment, including appropriate delivery systems and dosages of repair factors., Methods: The L5-L6 intervertebral disc was pierced with a 23-gauge needle in 18 rats. The animals received either sham or corticosterone-charged tricalcium phosphate ceramic capsules. The rats were euthanized at 4 weeks. Chondrocytes in the transition zone areas were counted and compared statistically., Results: The surgical technique induced degeneration of the nucleus without evidence of inflammation at adjacent levels when compared with nontraumatized controls. The number of chondrocytes per area was significantly less in the sham group than in the control group. Corticosteroid treatment showed chondrocyte numbers similar to control in 4 of 5 different views of the disc. The anterior region of the disc had 50% less chondrocytes per area than the control; however, the chondrocyte numbers were 50% greater than in the same site from discs of sham animals., Conclusions: The results show the development of a degenerative disc animal model that can be used to test the effects of growth enhancing factors in disc repair. Administration of continuous sustained release of corticosterone can slow the process of degeneration within the traumatized disc in the rat model.

Connective Tissue Growth Factor (CTGF) expression has been identified in a wide variety of fibrotic disorders; however, the expression of CTGF in carpal tunnel syndrome (CTS) has not yet been described in the literature. Both inflammatory and fibrotic etiologies have been implicated in the pathogenesis of CTS, with current evidence favoring an emphasis on a non-inflammatory fibrosis pathophysiological picture. Our objective was to identify whether CTGF is expressed in the tenosynovium of patients with CTS. Tenosynovial tissue was isolated from human subjects undergoing surgical decompression of the carpal tunnel (carpal tunnel release or CTR) for treatment of CTS following various durations of failed conservative management. Samples tested included patients with "idiopathic" CTS alone or CTS in the presence of associated co-orbidities including Type II Diabetes Mellitus (DM), Rheumatoid Arthritis (RA), and Systemic Lupus Erythematosus (SLE). SDS-PAGE protein analysis of tenosynovial tissue homogenate was performed to assess for differences in overall protein expression amongst all samples. Our findings demonstrate the presence varying levels of CTGF in tenosynovial samples from patients with CTS. Additionally, tenosynovial samples from patients with certain associated comorbidities - specifically RA and SLE - exhibit significant upregulation of CTGF levels relative to the levels observed patients with "idiopathic" CTS. These findings indicate that there is likely to be a role for CTGF in the pathogenesis of CTS.

Objective: Reliable platelet function monitoring is desirable in patients treated with glycoprotein IIb/IIIa receptor inhibitors. The aim of the present laboratory-based study was to assess platelet function after administration of clinically relevant doses of the glycoprotein IIb/IIIa antagonist tirofiban with or without heparin by using Sonoclot (Sienco Inc) and platelet aggregometry., Methods: Tirofiban (0-100 ng x mL(-1)) and heparin (0 or 1 U x mL(-1)) were added to blood samples obtained from 20 healthy volunteers. Coagulation analysis was performed on citrated whole blood by using the Sonoclot analyzer. The glass bead-activated test and the new glass bead test with heparinase were used. The results were compared with adenosine-5'-diphosphate-activated platelet aggregometry., Results: Administration of tirofiban showed a similar increase of platelet inhibition detected with the Sonoclot glass bead-activated test and glass bead test with heparinase, as well as by means of aggregometry. Bias between the different techniques was comparable; Spearman rank correlation was strong (glass bead-activated test vs aggregometry: rho = 0.823, P < .001; glass bead test with heparinase vs aggregometry: rho = 0.856, P < .001). After additional administration of heparin, platelet inhibition was only comparable for the glass bead test with heparinase and aggregometry, and the correlation coefficient remained unchanged for the glass bead test with heparinase versus aggregometry (rho = 0.878, P < .001). By contrast, the glass bead-activated test showed a nearly complete platelet inhibition with a significant bias compared with the glass bead test with heparinase and aggregometry. Correlation was weak for the glass bead-activated test versus aggregometry (rho = 0.407, P = .004)., Conclusions: When compared with platelet aggregometry, the glass bead-activated test from Sonoclot reliably detects glycoprotein IIb/IIIa receptor inhibition with tirofiban in unheparinized whole blood. However, in heparinized blood the glass bead test with heparinase is essential to accurately assess platelet function.

The objective of this study was to determine the effects of spaceflight on the structure of the tendon-bone junction (TBJ). Pregnant rats either flew in the space shuttle Atlantis (flight group; F) or were exposed to simulated launch and landing protocols (synchronous control group; SC) during days 9-19 of pregnancy. Following birth of their pups, maternal hindlimbs were studied using scanning electron and light microscopic histomophometric techniques. The tibial and calcaneal tuberosities, the fibular head, and the tibia-fibula junction were studied. Myofiber density and cross-sectional area of the quadratus femoris and soleus muscles and diameters of the calcaneal and patellar tendons were also evaluated. Cortical erosion was significantly greater at the tibial tuberosity and the fibular head in F animals compared to SC animals (P < 0.001). Sharpey fiber density was significantly less at the tibial tuberosity and fibular head in F animals compared to SC animals (P < 0.001). The myofiber area of both the soleus and quadratus femoris muscles and the diameters of both calcaneal and patellar tendons were significantly less in F compared to SC rats (P < 0.05). Our data illustrate that the TBJ morphology is affected by spaceflight at the attachment sites of the soleus and quadratus femoris muscles in pregnant animals, which could adversely affect their physical properties. These atrophic TBJ changes could have resulted from atrophy of the adjacent muscles and their tendons. Atrophic changes in the structure of the TBJ could predispose an animal to injury following spaceflight, when normal gravity conditions are reestablished.

Background: Antimitotic chemotherapeutic agents target tubulin, the major protein in mitotic spindles. Tubulin isotype composition is thought to be both diagnostic of tumor progression and a determinant of the cellular response to chemotherapy. This implies that there is a difference in isotype composition between normal and tumor tissues., Methods: To determine whether such a difference occurs in breast tissues, total tubulin was fractionated from lysates of paired normal and tumor breast tissues, and the amounts of beta-tubulin classes I + IV, II, and III were measured by competitive enzyme-linked immunosorbent assay (ELISA). Only primary tumor tissues, before chemotherapy, were examined. Her2/neu protein amplification occurs in about 30% of breast tumors and is considered a marker for poor prognosis. To gain insight into whether tubulin isotype levels might be correlated with prognosis, ELISAs were used to quantify Her2/neu protein levels in these tissues., Results: Beta-tubulin isotype distributions in normal and tumor breast tissues were similar. The most abundant beta-tubulin isotypes in these tissues were beta-tubulin classes II and I + IV. Her2/neu levels in tumor tissues were 5-30-fold those in normal tissues, although there was no correlation between the Her2/neu biomarker and tubulin isotype levels., Conclusion: These results suggest that tubulin isotype levels, alone or in combination with Her2/neu protein levels, might not be diagnostic of tumorigenesis in breast cancer. However, the presence of a broad distribution of these tubulin isotypes (for example, 40-75% beta-tubulin class II) in breast tissue, in conjunction with other factors, might still be relevant to disease progression and cellular response to antimitotic drugs.

Late aseptic loosening of total joint implants continues to be a common cause of implant failure. However, the pathophysiology of implant loosening remains controversial as to which factors at the implant tissue interface plays a crucial role in implant failure. The most prominent features of the foreign body membrane obtained from patients undergoing revision hip surgery were the presence of lymphocytes, histiocytes, giant cells, and immature collagen formation. The inflammatory sites are often characterized by the infiltration of activated lymphocytes and macrophages into the synovial membrane followed by proliferation of the synovial cells. The present study was conducted to determine if synovial cells in addition to macrophage cells are activated by tumor necrosis factor (TNF), which ultimately leads to foreign body membrane proliferation and ultimately joint space destruction. Macrophages or synovial cells were seeded at a density of 1 x 10(6) cells/ml and 5 ml were placed onto a 12 mm culture dish. The cells were challenged with either tissue culture media or media containing 2 ng/ml LPS or various doses of TNF (0.5, 5 and 50 ng/ml). The result indicated that both cell types were able to produce the inflammatory cytokines (IL-1 and IL-8 as early as 8 hours after a challenge with peak production occurring between 18-24 hours). The data suggest that both cell types are capable of eliciting inflammatory mediators, which can ultimately lead to joint destruction.

Ischemia-induced reperfusion injury seems to play an important role in the pathophysiology of "idiopathic" carpal tunnel syndrome (CTS). The common final pathway in this developmental sequence is thought to be an intermittent increase in interstitial pressure, leading to degenerative changes in the flexor tenosynovium and fibrotic changes in the perineural tissue. We hypothesize that this concurrently leads to alteration in the physical properties of the synovium, leading to its rapid and persistent swelling. A prospective study was conducted on synovial tissue obtained from 27 CTS patients. The in vitro synovial absorption rate of CTS patients was significantly higher in the first hour compared to controls (n = 7). This difference was maintained up to 5-6 h, albeit at a slower rate. Rapid absorption and retention of fluid by the synovium led to increased interstitial pressure and nerve compression, resulting in early and persistent manifestation of symptoms in sensitized patients., (Copyright 2002 Wiley-Liss, Inc. MICROSURGERY 22:316-319 2002)

In total, 41 consecutive patients with "idiopathic carpal tunnel syndrome" and abnormal electrophysiologic findings who underwent carpal tunnel release were studied prospectively. The focus of this investigation was the evaluation of the levels of specific chemical mediators within the serum and flexor tenosynovium of these patients. Blood was collected from these patients within 1 week prior to carpal tunnel release, and flexor tenosynovium was obtained at time of surgery. Specimens were then analyzed to determine the levels of interleukins 1 and 6, prostaglandin E(2) (PGE(2)), and malondialdehyde bis diethyl acetal. These values were compared to those of controls who had no evidence of carpal tunnel syndrome. A significant increase was noted in the serum malondialdehyde and tenosynovial levels of malondialdehyde, interleukin 6, and prostaglandin PGE(2) compared to controls. The elevated levels of these biologic factors and the absence of interleukin 1 elevation support a noninflammatory ischemia-reperfusion etiology for so-called "idiopathic carpal tunnel syndrome" that causes progressive edema and fibrosis of the tissues within the carpal canal. These findings correlate with previous histopathology reports. We believe that "idiopathic carpal tunnel syndrome" is an "-osis" not an "-itis.", (Copyright 2002 Wiley-Liss, Inc. MICROSURGERY 22:378-385 2002)