In vivo study of the efficacy of bupivacaine-eluting novel soy protein wound dressings in a rat burn model.

Dealing with wound related pain is an integral part of treatment. Systemic administration of analgesic and anesthetic agents is a common solution for providing pain relief to patients but comes at a risk of severe side effects as well as addiction. To overcome these issues, research efforts were madeto provide a platform for local controlled release of pain killers. We have developed a bilayer soy protein-based wound dressing for the controlled local release of bupivacaine to the wound site. The combination of a dense and a porous layer provides a platform for cell growth and proliferation as well as physical protection to the wound site. The current study focuses on the in vitro bupivacaine release profile from the dressing and the corresponding in vivo results of pain levels in a second-degree burn model on rats. The Rat Grimace Scale method and the Von Frey filaments method were used to quantify both, spontaneous pain and mechanically induced pain. A high burst release of 61.8 ± 1.9% of the loaded drug was obtained during the initial hour, followed by a slower release rate during the following day. The animal trials show that the RGS scores of the bupivacaine-treated group were significantly lower than these of the untreated group, proving a decrease of 51-68% in pain levels during days 1-3 after burn. Hence, successful pain reduction of spontaneous pain as well as mechanically induced pain, for at least three days after burn was achieved. It is concluded that our novel bupivacaine eluting soy protein wound dressings are a promising new concept in the field of local controlled drug release for pain management.

Strong bonding of corneal incisions using a noncontact fiber-optic laser soldering method.

Suturing of corneal incisions requires significant skill. We demonstrate a noncontact method that will simplify the bonding process. 5-mm-long penetrating vertical and slanted incisions were made in corneas of eyes, extracted from dead piglets. A fiber-optic laser system was used for laser soldering of the incisions, under close temperature control, using albumin solder. The burst-pressure PB immediately after the soldering was found to be PB ≈ 92 and 875 mmHg, for vertical and slanted incisions, respectively. PB = 875 mmHg is an exceptionally high figure, ≈10 times the clinically acceptable value for sutured incisions. Laser soldering was then performed on penetrating incisions made in the corneas of live healthy piglets, of weight ≈10 Kg. After a healing period, the eyes were extracted, and the corneas were examined by histopathology and by optical coherence tomography. Our method immediately generated watertight and strong bonding without noticeable corneal shape distortion. These results would be beneficial for cataract surgery and for corneal transplantations. The fiber-optic system makes it much easier to bond corneal incisions. In the future, laser soldering could be automated and efficiently used by less experienced surgeons, thereby reducing the workload on the experienced ones.

Subcutaneous Compared with Intraperitoneal KetamineXylazine for Anesthesia of Mice.

Mice are commonly anesthetized intraperitoneally with a ketamine-xylazine (KX) solution. Although this route of administration allows rapid uptake of the injected drugs, its disadvantages and potential risks include pain, peritoneal irritation, and perforation of an abdominal organ; some of the risks depend on the operator's experience. We compared the efficacy of intraperitoneal and subcutaneous administration of KX in HSD:ICR, BALB/cOlaHsd, and C57BL/6JOlaHsd mice in terms of time to onset and duration of surgical anesthesia, procedure safety, and mortality. Male and female mice (n = 20 each sex and strain) were anesthetized by using the same dose of intraperitoneal or subcutaneous KX. Time to onset and duration of immobilization and time to onset and duration of surgical anesthesia according to the pedal reflex differed significantly between strains. Within each strain, the durations of immobilization and surgical anesthesia were comparable between the routes of administration. The sex of the mouse but not the route of administration influenced whether surgical anesthesia was achieved. None of the subcutaneously-injected mice died. After intraperitoneal injections, 30% of the female mice died, compared with 3% of the male. In addition, fewer female mice achieved surgical anesthesia, suggesting a narrow therapeutic window for intraperitoneal KX in female mice. In conclusion, surgical anesthesia of mice with subcutaneous KX (K, 191.25 mg/kg; X, 4.25 mg/kg) seems to be safe, and the subcutaneous route is generally just as effective as the intraperitoneal route. The variability among mouse strains and between sexes requires further investigation to determine the optimal dosage.

Inadequate reinforcement of transmedial disruptions at branch points subtends aortic aneurysm formation in apolipoprotein-E-deficient mice.

INTRODUCTION: Infusion of angiotensin-II (Ang-II) in apolipoprotein-E-deficient mice (Apo-E(-/-)) results in suprarenal abdominal aortic aneurysm (AAA) in 30-85% of cases. This study identifies the apparent mechanism by which some animals do, but others do not, develop AAA in this model. METHODS: Male Apo-E(-/-) mice were infused with Ang-II (n=21) or saline (n=6) and sacrificed at 4 weeks. Aortas were excised, embedded in paraffin, sectioned (250 µm intervals), and stained. Sites of transmedial disruption (TMD) were identified and characterized, and their relationship to the 4 major aortic side branches (celiac, superior mesenteric, and renals) were determined. RESULTS: The frequency of TMDs in Ang-II-infused mice that formed AAA (n=9) was similar to those that did not (n=12) (AAA vs. no-AAA: 25 of 36[69%] vs. 28 of 48[58%] branches, P=.3 by chi-square). All TMDs were at branch points. However, in animals with AAA, the mean maximum length of the TMDs was significantly larger (1.94±1.6 vs. 0.65±0.5mm, P=.007 by Mann Whitney U test), the #mac-2(+) macrophages per 0.01mm(2) of defect area was greater (32±10 vs. 19±11, P<.02 by Kruskal-Wallis with Conover-Inman post hoc), the % area of attempted repair occupied by collagen was less (17±13% vs. 44±15%, P=.0009 by Mann Whitney U test), and the density of collagen per unit length of media missing was also markedly less (0.13±0.2 vs. 1.14±1.0, P=.0001 by Mann Whitney U test). CONCLUSIONS: Reinforcement of transmedial defects at branch points by wall matrix is a key intrinsic player in limiting AAA formation in the Ang-II-infused, Apo E(-/-) mouse and a potentially important mechanism-based therapeutic target for management of small, slowly progressing aneurysms.

Low level laser arrests abdominal aortic aneurysm by collagen matrix reinforcement in apolipoprotein E-deficient mice.

BACKGROUND AND OBJECTIVES: Recent in vitro studies by our group indicated that low level laser irradiation (LLLI) modifies cellular processes essential to the progression of abdominal aortic aneurysm (AAA). Using high-frequency ultrasonography (HF-u/s) in the angiotensin-II (Ang-II)-infused, apolipoprotein-E-deficient (Apo-E(-/-) ) mouse model of AAA, we found that LLLI markedly inhibited aneurysm formation and preserved arterial wall elasticity. We now report, using quantitative histopathology, the likely mechanism underlying the preventative effect of LLLI on aneurysm formation in this model. STUDY DESIGN/MATERIALS AND METHODS: This study was performed on 32 Apo-E(-/-) mice of which 10 were Ang-II-infused and LLL-irradiated (780 nm, 2 J/cm(2) , 9-minutes), 12 were Ang-II-infused but not irradiated, and 10 were saline infused. The aortas were excised at 28d, sectioned at 250 µm intervals, and stained with H + E, Movat-pentachrome and picrosirius-red for histomorphometry, and immunostained with Mac-2 and α-actin for detection of macrophages and SMCs, respectively. RESULTS: Transmural disruptions of the aorta occurred with distinct predilection for branch orifices. In the LLLI-treated animals, the frequency of these disruptions was lower (#branches with break points: 17 of 40 vs. 32 of 48, P = 0.023 by Chi-squared), their size smaller (length [mm]: 0.48 ± 0.26 vs. 0.98 ± 1.42, P = 0.044 by ANOVA with FPLSD), and the number of Mac-2-positive macrophages in the intramural areas of these disruptions lower than in the non-treated control (#Macrophages/0.01 mm(2) at break points: 11.6 ± 7.2 vs. 26.0 ± 15.7, P = 0.016 by Kruskal-Wallis). The average size of the medial SMCs was larger reflecting a heightened synthetic state (SMC size [µm(2) ]: 463.9 ± 61.4 vs. 354.9 ± 71.7, P = 0.001 by ANOVA with FPLSD). Furthermore, at sites of transmural disruption, the %area occupied by collagen of the overall area of attempted repair (%Col/WO) was significantly greater in the LLLI-treated animals versus control (%Col/WO: 41 ± 13 vs. 32 ± 16, P = 0.009 by ANOVA with FPLSD). CONCLUSION: Enhanced matrix reinforcement and modification of the inflammatory response at sites of transmural injury are prominent mechanisms by which LLLI reduces AAA progression in this model.

Cerebrospinal fluid segregation optic neuropathy: an experimental model and a hypothesis.

AIM: To describe the histological changes in the optic nerve (ON) after experimental segregation of cerebrospinal fluid (CSF). METHODS: In seven sheep, a silicone band was placed around one ON to compress the subarachnoid space (SAS) surrounding the nerve, thus blocking the flow of CSF without compressing the ON itself. After 4 or 21 days, both the ligated and untouched ONs were removed and evaluated histologically. RESULTS: All treated ONs showed marked loss of axons, destruction of myelin and swelling of meningoepithelial cells, most pronounced in the proximal ON adjacent to the globe at the location most distant to the ligature. There was no significant difference in histological findings between the ONs that were ligated for 4 days and those with 21 days of ligature. CONCLUSION: CSF segregation in the ON by blocking the SAS leads within 4 days to severe nerve damage. The increasing severity of these changes with increasing distance from the site of the ligature argues against simple pressure- or microperfusion-dependent effects and supports the hypothesis that interruption of CSF flow in the SAS of the ON can produce damage due to a change of CSF flow and content.

Low-level laser irradiation inhibits abdominal aortic aneurysm progression in apolipoprotein E-deficient mice.

AIMS: Increased early detection of abdominal aortic aneurysm (AAA) and the severe complications of its current treatment have emphasized the need for alternative therapeutic strategies that target pathogenetic mechanisms of progression and rupture. Recent in vitro studies from our laboratory have shown that low-level laser irradiation (LLLI) (780 nm) modifies cellular processes fundamental to aneurysm progression. The present study was designed to determine whether LLLI retards the progression of suprarenal AAA in vivo. METHODS AND RESULTS: High-frequency ultrasonography (0.01 mm resolution) was used to quantify the effect of LLLI on aneurysmatic aortic dilatation from baseline to 4 weeks after subcutaneous infusion of angiotensin II by osmotic minipumps in the apolipoprotein E-deficient mouse. At 4 weeks, seven of 15 non-irradiated, but none of the 13 LLLI, mice had aneurysmal dilatation in the suprarenal aneurysm-prone segments that had progressed to >or=50% increase in maximal cross-sectional diameter (CSD) over baseline (P = 0.005 by Fisher's exact test). The mean CSD of the suprarenal segments (normalized individually to inter-renal control segments) was also significantly lower in irradiated animals (LLLI vs. non-irradiated: 1.32 +/- 0.14 vs. 1.82 +/- 0.39, P = 0.0002 by unpaired, two-tailed t-test) with a 94% reduction in CSD at 4 weeks compared with baseline. M-mode ultrasound data showed that reduced radial wall velocity seen in non-treated was significantly attenuated in the LLLI mice, suggesting a substantial effect on arterial wall elasticity. CONCLUSION: These in vivo studies, together with previous in vitro studies from this laboratory, appear to provide strong evidence in support of a role for LLLI in the attenuation of aneurysm progression. Further studies in large animals would appear to be the next step towards testing the applicability of this technology to the human interventional setting.