Transduced PEP-1-FK506BP ameliorates corneal injury in Botulinum toxin A-induced dry eye mouse model

FK506 binding protein 12 (FK506BP) belongs to a family of immunophilins, and is involved in multiple biological processes. However, the function of FK506BP in corneal disease remains unclear. In this study, we examined the protective effects on dry eye disease in a Botulinum toxin A (BTX-A) induced mouse model, using a cell-permeable PEP-1-FK506BP protein. PEP-1-FK506BP efficiently transduced into human corneal epithelial cells in a time- and dose-dependent manner, and remained stable in the cells for 48 h. In addition, we demonstrated that topical application of PEP-1-FK506BP was transduced into mouse cornea and conjunctiva by immunohistochemistry. Furthermore, topical application of PEP-1-FK506BP to BTX-A-induced mouse model markedly inhibited expression levels of pro-inflammatory cytokines such as interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and macrophage inhibitory factor (MIF) in corneal and conjunctival epithelium. These results suggest PEP-1-FK506BP as a potential therapeutic agent for dry eye diseases. [BMB Reports 2013; 46(2): 124-129]     

PEP-1-FK506BP12 inhibits matrix metalloproteinase expression in human articular chondrocytes and in a mouse carrageenan-induced arthritis model

The 12 kDa FK506-binding protein (FK506BP12), an immunosuppressor, modulates T cell activation via calcineurin inhibition. In this study, we investigated the ability of PEP-1-FK506BP12, consisting of FK506BP12 fused to the protein transduction domain PEP-1 peptide, to suppress catabolic responses in primary human chondrocytes and in a mouse carrageenan-induced paw arthritis model. Western blotting and immunofluorescence analysis showed that PEP-1-FK506BP12 efficiently penetrated chondrocytes and cartilage explants. In interleukin-1β (IL-1β)-treated chondrocytes, PEP-1-FK506BP12 significantly suppressed the expression of catabolic enzymes, including matrix metalloproteinases (MMPs)-1, -3, and -13 in addition to cyclooxygenase-2, at both the mRNA and protein levels, whereas FK506BP12 alone did not. In addition, PEP-1-FK506BP12 decreased IL-1β-induced phosphorylation of the mitogen-activated protein kinase (MAPK) complex (p38, JNK, and ERK) and the inhibitor kappa B alpha. In the mouse model of carrageenan-induced paw arthritis, PEP-1-FK506BP12 suppressed both carrageenan-induced MMP-13 production and paw inflammation. PEP-1-FK506BP12 may have therapeutic potential in the alleviation of OA progression. [BMB Reports 2015; 48(7): 407-412]     

PEP-1-FK506BP inhibits alkali burn-induced corneal inflammation on the rat model of corneal alkali injury

FK506 binding protein 12 (FK506BP) is a small peptide with a single FK506BP domain that is involved in suppression of immune response and reactive oxygen species. FK506BP has emerged as a potential drug target for several inflammatory diseases. Here, we examined the protective effects of directly applied cell permeable FK506BP (PEP-1-FK506BP) on corneal alkali burn injury (CAI). In the cornea, there was a significant decrease in the number of cells expressing pro-inflammation, apoptotic, and angiogenic factors such as TNF-α, COX-2, and VEGF. Both corneal opacity and corneal neovascularization (CNV) were significantly decreased in the PEP-1-FK506BP treated group. Our results showed that PEP-1-FK506BP can significantly inhibit alkali burn-induced corneal inflammation in rats, possibly by accelerating corneal wound healing and by reducing the production of angiogenic factors and inflammatory cytokines. These results suggest that PEP-1-FK506BP may be a potential therapeutic agent for CAI. [BMB Reports 2015; 48(11): 618-623]     

Fenobam promoted the neuroprotective effect of PEP-1-FK506BP following oxidative stress by increasing its transduction efficiency

We examined the ways in which fenobam could promote not only the transduction of PEP-1-FK506BP into cells and tissues but also the neuroprotective effect of PEP-1-FK506BP against ischemic damage. Fenobam strongly enhanced the protective effect of PEP-1-FK506BP against H₂O₂-induced toxicity and DNA fragmentation in C6 cells. In addition, combinational treatment of fenobam with PEP-1-FK506BP significantly inhibited the activation of Akt and MAPK induced by H₂O₂, compared to treatment with PEP-1-FK506BP alone. Interestingly, our results showed that fenobam significantly increased the transduction of PEP-1-FK506BP into both C6 cells and the hippocampus of gerbil brains. Subsequently, a transient ischemic gerbil model study demonstrated that fenobam pretreatment led to the increased neuroprotection of PEP-1-FK506BP in the CA1 region of the hippocampus. Therefore, these results suggest that fenobam can be a useful agent to enhance the transduction of therapeutic PEP-1-fusion proteins into cells and tissues, thereby promoting their neuroprotective effects. [BMB Reports 2013; 46(11): 561-566]     

PEP-1-HO-1 prevents MPTP-induced degeneration of dopaminergic neurons in a Parkinson’s disease mouse model

Heme oxygenase-1 (HO-1) degrades heme to carbon dioxide, biliverdin, and Fe²⁺, which play important roles in various biochemical processes. In this study, we examined the protective function of HO-1 against oxidative stress in SH-SY5Y cells and in a Parkinson’s disease mouse model. Western blot and fluorescence microscopy analysis demonstrated that PEP-1-HO-1, fused with a PEP-1 peptide can cross the cellular membranes of human neuroblastoma SH-SY5Y cells. In addition, the transduced PEP-1-HO-1 inhibited generation of reactive oxygen species (ROS) and cell death caused by 1-methyl-4-phenylpyridinium ion (MPP⁺). In contrast, HO-1, which has no ability to transduce into SH-SY5Y cells, failed to reduce MPP⁺-induced cellular toxicity and ROS production. Furthermore, intraperitoneal injected PEP-1-HO-1 crossed the blood-brain barrier in mouse brains. In a PD mouse model, PEP-1-HO-1 significantly protected against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced toxicity and dopaminergic neuronal death. Therefore, PEP-1-HO-1 could be a useful agent in treating oxidative stress induced ailments including PD. [BMB Reports 2014; 47(10): 569-574]     

Quantitative assessment of tear production: A review of methods and utility in dry eye drug discovery

The successful development of a therapeutic agent targeting treatment of dry eye syndrome necessitates the demonstration of drug efficacy for both sign and symptom endpoints. As numerous therapeutic strategies incorporate a secretagogue function into their overall mechanism of action, the quantitative assessment of tear production serves as a logical endpoint to anchor “sign” efficacy. Although several methods including the Schirmer, the phenol red thread and tear clearance tests exist, their utility in clinical evaluations of novel therapeutics is unclear. The purpose of this review is to summarize findings and conclusions describing the performance of each of these tests so as to gain insight into which, if any, is most applicable for use in discovering new dry eye therapeutics.     

Cytotoxic effects of FK506 on human renal proximal tubule cells in culture

Summary FK506 has been used as the primary immunosuppressive agent administered after a variety of organ transplants, with less reported nephrotoxicity than that of cyclosporine. This study examined in vitro cytotoxicity of FK506 on normal human renal proximal tubule cells. Cytotoxicity was assessed by neutral red inclusion and trypan blue exclusion; morphology was assessed by light and transmission electron microscopy. Neutral red inclusion decreased to less than 10% of the control after 3 days exposure to 200μg/ml FK506. Forty microgram per milliliter FK506 caused a decrease in neutral red inclusion to 61% of the control on Day 7, with recovery to 86% on Day 12. Similarly, trypan blue exclusion decreased to 66% of the control following 7 days exposure to 40μg/ml FK506, and confluency of the monolayer was reduced to 50% as evidenced by phase contrast microscopy. After a 12-day exposure, treated monolayers became more confluent. On ultrastructural examination, FK506-treated cells exhibited increased cytoplasmic vacuolation and lipid inclusion. These data suggest that FK506 is reversibly and mildly toxic to monolayers of human renal proximal tubule cells and are consistent with clinical reports of reversible nephrotoxicity.     

Beneficial effect of daidzin in dry eye rat model through the suppression of inflammation and oxidative stress in the cornea

Present investigation evaluates the effect of daidzin in dry eye rat model through the suppression of inflammation and oxidative stress in the cornea. Briefly, electron spine resonance was used for the estimation of radical scavenging activity of daidzin and COX Fluorescent Activity Assay Kit was used for the estimation of PGS activity. Dry eye rat model was developed by removing the lacrimal gland and effect of daidzin was evaluated in dry eye rat model by estimating the fluorescein score, tear volume and expressions of heme oxigenase (HO-1), TNF α, Interlukin 6 (IL-6), matrix metallopeptidase 9 (MMP-9) and PGS-2. Result of the present study suggested that daidzin possess tyrosyl radical scavenging activity and thereby decreases the oxidative stress. Activity of PGS significantly increases in dry eye which was inhibited by daidzin treatment due to competitive inhibition of PGS. It also recovers the tear volume in dry eye rat model in which lacrimal gland was removed. Thus corneal erosion was improved by daidzin in dry eye rat model. Thus present study concludes that treatment with daidzin protects the cornea in dry eye rat model by suppression inflammation and oxidative stress.     

Anti-inflammatory effect of transduced PEP-1-heme oxygenase-1 in Raw 264.7 cells and a mouse edema model

Heme oxygenase-1 (HO-1), which catalyzes the degradation of free heme to biliverdin, carbon monoxide (CO), and free iron (Fe²⁺), is up-regulated by several cellular stress and cell injuries, including inflammation, ischemia and hypoxia. In this study, we examined whether fusion of HO-1 with PEP-1, a protein transduction domain that is able to deliver exogenous molecules to living cells or tissues, would facilitate HO-1 delivery to target cells and tissues, and thereby effectively exert a therapeutically useful response against inflammation. Western blot analysis demonstrated that PEP-1-HO-1 fusion proteins were transduced into Raw 264.7 cells in time- and dose-dependent manners, and were stably maintained in the cells for about 60 h. In addition, fluorescence analysis revealed that only PEP-1-HO-1 fusion proteins were significantly transduced into the cytoplasm of cells, while HO-1 proteins failed to be transduced. In lipopolysaccharide (LPS)-stimulated Raw 264.7 cells and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced mouse edema model, transduced PEP-1-HO-1 fusion proteins effectively inhibited the overexpression of pro-inflammatory mediators and cytokines. Also, histological analysis demonstrated that PEP-1-HO-1 remarkably suppressed ear edema. The results suggest that the PEP-1-HO-1 fusion protein can be used as a therapeutic molecule against reactive oxygen species-related inflammatory diseases.     

Proteomic analysis revealed the altered tear protein profile in a rabbit model of Sjögren's syndrome‐associated dry eye

Sjögren's syndrome (SS) is an autoimmune disease that results in pathological dryness of mouth and eye. The diagnosis of SS depends on both clinical evaluation and specific antibodies. The goal of this study was to use quantitative proteomics to investigate changes in tear proteins in a rabbit model of SS‐associated dry eye, induced autoimmune dacryoadenitis (IAD). Proteomic analysis was performed by iTRAQ and nano LC‐MS/MS on tears collected from the ocular surface, and specific proteins were verified by high resolution MRM. It was found that in the tears of IAD rabbits at 2 and 4 weeks after induction, S100 A6, S100 A9, and serum albumin were upregulated, whereas serotransferrin (TF), prolactin‐inducible protein (PIP), polymeric immunoglobulin receptor (pIgR), and Ig gamma chain C region were downregulated. High resolution MRM with mTRAQ labeling verified the changes in S100 A6, TF, PIP, and pIgR. Our results indicated significant changes of tear proteins in IAD rabbits, suggesting these proteins could potentially be used as biomarkers for the diagnosis and prognosis of dry eye. Several of these proteins were also found in the tears of non‐SS dry eye patients indicating a common basis of ocular surface pathology, however, pIgR appears to be unique to SS.     

Punctal plugs versus artificial tears for treating dry eye: a comparative observation of their effects on contrast sensitivity

This study aimed to compare the effects of treatment with punctal plugs versus artificial tears on visual function and tear film stability for dry eye. A total of 56 consecutive eyes of 28 dry eye patients observed at our clinic from May to October in 2009 were divided into two groups. One group (32 eyes of 16 patients) was treated with artificial tears, and punctal plugs were used in the other group (24 eyes of 12 patients). A questionnaire was used in these patients before treatment and was repeated 2 weeks after treatment. Fluorescent staining for tear film break-up time (BUT), the Schirmer test I (STI), and contrast sensitivity was performed at the same time. The questionnaire indicated that all patients complained about the uncomfortable symptoms associated with dry eye. These symptoms were relieved after the application of artificial tears or punctal plugs, and there was no significant difference between these two groups. We found that the corneal fluorescent staining disappeared after treatment. The BUT was improved significantly after treatment in both groups, but the improvement was greater in patients who received punctal plugs than those that received artificial tears. There was no remarkable change in the STI in the artificial tears group, but a significant change was observed in the punctal plugs group. The contrast sensitivities were greatly improved in simulated daylight, night, and glare disability conditions after treatment with artificial tears and punctal plugs. However, the changes in contrast sensitivity did not significantly differ between groups. Both artificial tears and punctal plugs relieved dry eye symptoms, repaired corneal lesions, enhanced tear film stability, and improved contrast sensitivity. Punctal plugs could improve tear film stability and elongate the BUT better than artificial tears.     

A comparative study on immunosuppressive effects of cyclosporin A and FK 506 on peripheral blood lymphocytes in dogs

Immunosuppressive effects of cyclosporin A (CsA) and FK 506 (FK) on peripheral blood lymphocytes were studied in dogs in respect to mixed lymphocyte reaction, proliferative responses to recombinant interleukin-2 (rIL-2), phytohemagglutinin (PHA) and concanavalin-A (Con-A); phenotypes of OKIa1, CD3, CD8 and surface IgM; cytotoxic activity against xenogeneic tumor cells. CsA (2.0 or 5.0 mg/kg, intravenously) or FK (0.16 mg/kg, intramuscularly) was given to mongrel dogs every morning for serial 21 days. The blood concentrations of CsA, measured as trough levels by fluorescence polarization method, ranged from 37 to 350 ng/ml in dogs administered at 2.0 mg/kg and from 170 to 894 ng/ml in dogs administered at 5.0 mg/kg during treatment, respectively. In dogs treated with FK at a dose of 0.16 mg/kg, the drug concentrations in the plasma during treatment ranged from 0.16 to 1.8 ng/ml. Mixed lymphocyte reaction and proliferative responses to rIL-2, PHA and Con-A, which were declined by CsA, were not affected by FK. In contrast, the proportion of OKIa1+ cells was not affected by CsA, whereas FK decreased the proportion of OKIa1+ cells progressively during the course of treatment. Cytotoxic activity was suppressed by both CsA and FK. These results possibly indicate that CsA and FK exert their immunosuppressive effects via different mechanisms.     

Neuroprotective effect of PEP-1-peroxiredoxin2 on CA1 regions in the hippocampus against ischemic insult

Background

Oxidative stress is a leading cause of various diseases, including ischemia and inflammation. Peroxiredoxin2 (PRX2) is one of six mammalian isoenzymes (PRX1–6) that can reduce hydrogen peroxide (H₂O₂) and organic hydroperoxides to water and alcohols.

Methods

We produced PEP-1-PRX2 transduction domain (PTD)-fused protein and investigated the effect of PEP-1-PRX2 on oxidative stress-induced neuronal cell death by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Western blot, immunofluorescence microscopy, and immunohistochemical analysis.

Results

Our data showed that PEP-1-PRX2, which can effectively transduce into various types of cells and brain tissues, could be implicated in suppressing generation of reactive oxygen species, preventing depolarization of the mitochondrial membrane, and inhibiting the apoptosis pathway in H₂O₂-stimulated HT22, murine hippocampal neuronal cells, likely resulting in protection of HT22 cells against H₂O₂-induced toxicity. In addition, we found that in a transient forebrain ischemia model, PEP-1-PRX2 inhibited the activation of astrocytes and microglia in the CA1 region of the hippocampus and lipid peroxidation and also prevented neuronal cell death against ischemic damage.

Conclusions

These findings suggest that the transduced PEP-1-PRX2 has neuroprotective functions against oxidative stress-induced cell death in vitro and in vivo.

General significance

PEP-1-PRX2 could be a potential therapeutic agent for oxidative stress-induced brain diseases such as ischemia.     

Longitudinal analysis of taurine induced effects on the tear proteome of contact lens wearers and dry eye patients using a RP-RP-Capillary-HPLC-MALDI TOF/TOF MS approach

Tear proteomic studies revealed distinct similarities between contact lens wearers and dry eye patients. AMO Complete® multipurpose contact lens cleaning solutions containing taurine seem to have a beneficial effect regarding contact lens induced dry eye. To illuminate the effect of taurine on the tear proteome of contact lens wearers and sicca patients we developed a gel-based RP-RP capillary HPLC–MALDI TOF/TOF MS strategy. Two contact lens wearer groups, one using eye drops containing 0.05% taurine; the other for control physiological NaCl solution were monitored. Also, a third group of sicca patients using taurine solution was studied (N = 4 individuals/group). Tear pools of each group at six time points over 5 weeks were analyzed. In summary 267 tear proteins were identified. We found a protein subset showing a linear taurine response with R² values ≥ 0.5. Taurine effects were detected predominantly in the contact lens group demonstrated by distinct level decreases. Most protein candidates were related to inflammation. Since levels of these proteins differentiate from those of a healthy non-contact lens wearer reference they are supposed to be involved in contact lens induced dry eye and should be focused on in further studies.     

Focused ultrasound stimulation on meibomian glands for the treatment of evaporative dry eye

Current treatments for meibomian gland dysfunction have several limitations, creating a necessity for other advanced treatment options. The purpose of this study is to determine the effectiveness of focused ultrasound stimulation for the treatment of dry eye disease caused by meibomian gland dysfunction. An in vivo study of nine Dutch Belted rabbits was conducted with focused ultrasound stimulation of the meibomian glands. A customized line-focused ultrasonic transducer was designed for treatment. Fluorescein imaging, Schirmer’s test, and Lipiview II ocular interferometer were used to quantify outcomes from three aspects: safety, tear production, and lipid layer thickness. Both tear secretion and lipid layer thickness improved following ultrasound treatment. Five to 10 min after the ultrasound treatment, the mean values of lipid layer thickness increased from 55.33 ± 11.15 nm to 95.67 ± 22.77 nm (p < 0.05), while the mean values measured with the Schirmer’s test increased from 2.0 ± 2.3 to 7.2 ± 4.3 (p < 0.05). Positive effects lasted more than three weeks. Adverse events such as redness, swelling, and mild burn, occurred in two rabbits in preliminary experiments when the eyelids sustained a temperature higher than 42°C. No serious adverse events were found. The results suggest that ultrasound stimulation of meibomian glands can improve both tear production and lipid secretion. Ultimately, ultrasound stimulation has the potential to be an option for the treatment of evaporative dry eye disease caused by meibomian gland dysfunction.     

Dose-dependent effects of FK506 on neuroregeneration in a rat model

This study explored the effects of different doses of FK506 on peripheral nerve regeneration, to determine whether neuroregeneration could be enhanced without the toxicity of systemic immunosuppression. In the first part of the study, subimmunosuppressive doses of FK506 were determined by examining skin allograft survival in a rat model. Full-thickness skin grafts (2 cm2) from Wistar rats were grafted to recipient Lewis rats. The procedure was performed for six groups (n = 6). The control group received no FK506, and the other five groups received daily doses of FK506 of 0.125, 0.25, 0.5, 1.0, or 2.0 mg/kg. Animals that received 2.0 mg/kg FK506 per day exhibited complete skin graft take, whereas all other groups demonstrated complete rejection. After determination of the immunosuppressive dose of FK506, the neuroregenerative effects of different doses of FK506 were explored by assessing nerve regeneration in 80 rats after tibial nerve transection and repair. The control group received no FK506, whereas the other four groups were given daily doses of FK506 of 0.25, 0.5, 1.0, or 2.0 mg/kg. Rats were euthanized at three time points (25, 30, and 35 days), to fully investigate the effects of different FK506 dosing regimens on neuroregeneration. Histomorphometric analyses performed on postoperative days 30 and 35 demonstrated statistically significant improvements in neuroregeneration with subimmunosuppressive FK506 doses of 0.5 and 1.0 mg/kg per day. Therefore, the study demonstrated that neuroregeneration was enhanced at low doses of FK506 that were not sufficient to prevent skin allograft rejection.     

PEP-1-PEA-15 protects against toxin-induced neuronal damage in a mouse model of Parkinson's disease

Background

PEA-15 is abundantly expressed in both neurons and astrocytes throughout the brain. It is a multifunctional protein with the ability to increase cell survival via anti-apoptotic and anti-proliferative properties. However, the function of PEA-15 in neuronal diseases such as Parkinson's disease (PD) remains unclear. In this study, we investigated the protective effects of PEA-15 on neuronal damage induced by MPP⁺ in neuroblastoma SH-SY5Y and BV2 microglia cells and in a MPTP-induced PD mouse model using cell-permeable PEP-1-PEA-15.

Methods

PEP-1-PEA-15 was purified using affinity chromatography. Cell viability and DNA fragmentation were examined by MTT assay and TUNEL staining. Dopaminergic neuronal cell death in the animal model was examined by immunohistochemistry.

Results

PEP-1-PEA-15 transduced into the SH-SY5Y and BV2 cells in a time- and dose-dependent manner. Transduced PEP-1-PEA-15 protected against MPP⁺-induced toxicity by inhibiting intracellular ROS levels and DNA fragmentation. Further, it enhanced the expression levels of Bcl-2 and caspase-3 while reducing the expression levels of Bax and cleaved caspase-3. We found that PEP-1-PEA-15 transduced into the substantia nigra and prevented dopaminergic neuronal cell death in a MPTP-induced PD mouse. Also, we showed the neuroprotective effects in the model by demonstrating that treatment with PEP-1-PEA-15 ameliorated MPTP-induced behavioral dysfunctions and increased dopamine levels in the striatum.

Conclusions

PEP-1-PEA-15 can efficiently transduce into cells and protects against neurotoxin-induced neuronal cell death in vitro and in vivo.

General significance

These results demonstrate the potential for PEP-1-PEA-15 to provide a new strategy for protein therapy treatment of a variety of neurodegenerative diseases including PD.     

Tat-Frataxin protects dopaminergic neuronal cells against MPTP-induced toxicity in a mouse model of Parkinson's disease

Parkinson's disease (PD) is caused by various factors such as reactive oxygen species (ROS), dysfunction of mitochondria, and aggregation of misfolded proteins, thereby leading to loss of dopaminergic (DA) neurons in the substantia nigra (SN) of the brain. Frataxin (FXN) is associated with iron homeostasis and biogenesis of iron-sulfur clusters in the electron transport chain complex. In this study, we investigated the potential of Tat-FXN to cross the blood-brain barrier (BBB) and protect DA neurons against oxidative stress in a mouse model of PD. Tat-FXN was effectively transduced into SH-SY5Y cells and blocked production of ROS and cleavage of DNA, significantly improving cell survival against 1-methyl-4-phenylpyridinium induced toxicity. In addition, Tat-FXN efficiently penetrated the BBB and exhibited a clear neuroprotective effect on tyrosine hydroxylase-specific DA neurons in the SN in a mice model of 1-methyl-4-phenyl-1,2,3,6-tetra-hydropyridine-induced PD. Therefore, these results suggest that Tat-FXN may provide neuroprotective therapy for ROS related diseases including PD.     

Antimicrobial and bone repair effects of boric acid in a rat model of dry socket (alveolar osteitis) following dental extraction

BackgroundAlveolitis occurs after dental extraction without blood clot formation, leading to an inflammatory process and bacterial contamination. Boric acid (BA) demonstrates anti-inflammatory, antimicrobial, and osteogenic properties. This study aims to evaluate the possible antimicrobial effects and bone repair of BA in a rat model of alveolitis (dry socket).Methods33 male Wistar rats were submitted to the extraction of the upper right incisor and dry socket induction. They were first divided into two groups: dry socket (n = 17) and dry socket + 0.75 % BA (n = 16). Samples for the microbiological analysis were collected immediately after dental extraction, at the detection of clinical alveolitis, 7, and 14 days after BA application. For microCT and histological analysis, samples from euthanized rats were used in 14 and 28 days after alveolitis detection.ResultsHigher bacterial counts were found in 4–5 days after alveolitis induction, compared to the baseline in both experimental groups, decreasing significantly after 7 and 14 days of treatment with BA (P < 0.05). The microCT evaluation displayed increased bone volume, bone volume fraction, trabecular thickness, and bone mineral density in a time-dependent manner, regardless of BA treatment. On the other hand, the number of trabeculae and total bone porosity decreased over the 28 days of the experiment in the dry-socket group and both groups, respectively (P < 0.05). Histological analysis did not differ on bone repair in both experimental groups.ConclusionThis was the first report investigating the effects of BA in a rat model of alveolitis regarding microbiological and bone repair aspects. The BA local application decreased the total aerobic and facultative bacteria counts and does not seem to benefit the bone repair after alveolitis development. This study paves the way for more studies involving alveolitis and different BA applications.