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The recent change from the popular carboxamide to an acetamide (ATA) linker scaffold in synthetic cannabinoid receptor agonists (SCRAs) can be interpreted as an attempt to circumvent legal regulations, setting new analytical challenges. Metabolites of N-cyclohexyl-2-(1-pentyl-1 H-indol-3-yl)acetamide: CH-PIATA, the second ATA type SCRA detected in the EU, were investigated in urine and serum samples by LC-HRMS-MS and LC-MS-MS. Two different in vitro models, a pHLM assay and HepG2-cells, as well as an in silico prediction by GLORYx freeware assisted in metabolite formation/identification. CH-PIATA was extensively metabolized, leading to metabolites formed primarily by mono- and dihydroxylation. For urine and serum specimens, monohydroxylation at the indole core or the methylene spacer of the acetamide linker (M1.8), carboxylic acid formation at the N-pentyl side chain (M3.1) and degradation of the latter leading to a tentatively identified N-propionic acid metabolite (M5.1) are suggested as reliable markers for substance intake. The N-propionic acid metabolite could not be confirmed in the in vitro assays as it includes multiple consecutive metabolic reactions. Furthermore, CH-PIATA could be detected as parent substance in blood samples, but not in urine. Both in vitro assays and the in silico tool proved suitable for predicting metabolites of CH-PIATA. Considering effort and costs, pHLM incubations seem to be more effective for metabolite prediction in forensic toxicology than HepG2 cells. The highlighted Phase I metabolites serve as reliable urinary targets for confirming CH-PIATA use. The in silico approach is advantageous when reference material is unavailable., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site–for further information please contact journals.permissions@oup.com.)

Synthetic cannabinoid receptor agonists (SCRAs) are one of the largest groups of new psychoactive substances (NPS). Yet, another novel analog started spreading on the NPS market around 2021. Soon after, the substance could be analytically characterized in herbal material as ADB-HEXINACA, an SCRA containing a hexyl-substituted tail on the indazole core. Here, we present suitable urinary markers to prove the consumption of this analog, a case report of acute polydrug intoxication and data on its prevalence in Germany. Anticipated phase I metabolites were detected in 12 authentic urine samples that were collected for abstinence control and analyzed by ultra-performance liquid chromatography coupled to a time-of-flight mass spectrometer (UPLC-qToF-MS). The results of in vivo samples were confirmed by analysis of in vitro incubates with pooled human liver microsomes (pHLMs). Forensic samples were used to assess the prevalence of ADB-HEXINACA. Thirty-two phase I metabolites were detected in the authentic urine samples. The main metabolites resulted from amide hydrolysis in combination with either monohydroxylation or ketone formation at the hexyl moiety (M15 and M26), the monitoring of which is recommended as a proof of consumption. ADB-HEXINACA was detected in 3.5% of SCRA positive urine samples collected for abstinence control in Freiburg up to December 2022 and in 5.5% of the SCRA positive blood/serum samples. The hexyl substituent of ADB-HEXINACA allows for the detection of specific urinary biomarkers suggested as analytical targets to confirm its prior intake. ADB-HEXINACA had a rather low prevalence in Germany, alternating months of higher prevalence with periods of total absence., (© 2024 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

Synthetic cannabinoid receptor agonists (SCRAs) are distributed on the drug market to produce THC-like effects while evading routine drug testing and legislation. The cyclobutylmethyl (CBM) and norbornylmethyl (NBM) side chain specifically circumvented the German legislation and led to the emergence of exploratory SCRAs in 2019-2021. The NBM SCRAs were detected post-amendment of the new psychoactive substances act in 2020, which scheduled all CBM SCRAs. All six SCRAs are full agonists at the cannabinoid receptor 1 compared with Δ 9 -tetrahydrocannabinol and CP-55,940. The CBM SCRAs showed binding affinities of K i : 29.4-0.65 nm and potencies of EC 50 : 483-40.1 nm (CBMICA << CBMINACA < CBMeGaClone). The norbornyl derivatives exhibited high affinities (K i : 1.87-0.25 nm), with indazole being the most affine. Functional activity data confirmed that the indazole derivative tends to be the most potent of all three NBM SCRAs (EC 50 : 169-1.78 nm). The sterically demanding NBM side chain increased the affinity and activity of almost all core structures. Future studies should be conducted on similarly voluminous side chain moieties. The 'life cycle' of all SCRAs on the drug market was less than a year. Notably, Cumyl-CBMICA was the most prevalent while also having the weakest cannabimimetic properties. Quantification of Cumyl-CBMICA during peak consumption in late 2019 and early 2020 revealed an increase in the concentration on the herbal material, which, together with forum entries and blog posts, corroborates the low in vitro cannabimimetic properties. Seizure prevalence data indicate that almost all SCRAs continue to be identified in 2022, potentially due to remaining stocks., (© 2022 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

In this work, we present evidence for the formation of transient stalks in aligned multilamellar stacks of lipid membranes. Just above the phase transition from the fluid ([Formula: see text]) lamellar phase to the rhombohedral phase (R), where lipid stalks crystallize on a super-lattice within the lipid bilayer stack, we observe a characteristic scattering pattern, which can be attributed to a correlated fluid of transient stalks. Excess (off-axis) diffuse scattering with a broad modulation around the position which later transforms to a sharp peak of the rhombohedral lattice, gives evidence for the stalk fluid forming as a pre-critical effect, reminiscent of critical phenomena in the vicinity of second-order phase transitions. Using high-resolution off-specular X-ray scattering and lineshape analysis we show that this pre-critical regime is accompanied by an anomalous elasticity behavior of the membrane stack, in particular an increase in inter-bilayer compressibility, i.e., a decrease in the compression modulus.

Purpose: This study aims to identify multiple ligament knee injury patterns that possess a high-risk of vascular lesion., Methods: We retrospectively compared torn ligament patterns and the presence of vascular lesions confirmed by magnetic resonance imaging and computed tomography angiography from 122 consecutive patients with diagnoses of multiple ligament knee injury made at the emergency department between January 2012 and December 2017. Patients were not eligible if they had an ipsilateral lower extremity lesion (dislocations or fractures at another level), initial evaluation at another hospital, or follow-up for less than 12 months. The primary outcomes were the comparison between the imaging findings of torn structures patterns and the presence of a vascular lesion., Results: We identified 48 eligible patients (50 knees) with multiligamentary knee lesions, of whom eight had popliteal artery damage, yielding an incidence of 16%. Our clinical examination detected six of these patients that were classified, according to the Schenck system, as KD-IIIL (6 knees) and KD-IIIM (2 knees). The odds of having a popliteal artery injury is 4.69 to 1 with a KD-IIIL injury that with any other type of injury on that classification (95% CI 0.960-22.98)., Conclusions: This data suggests that varus forces causing enough energy to produce a KD-IIIL lesion possess a higher popliteal artery injury risk, making recommendable a thorough examination of the vascular integrity when diagnosing a KD-IIIL lesion.

Impact Statement: The promoter characterized in this study has been made accessible as a resource for the skeletal tissue engineering and regenerative medicine community. When combined with suitable reporter vectors, the resulting tools can be used for noninvasive and/or high-throughput screening of test conditions for stimulating chondrogenesis by candidate stem/progenitor cells. As demonstrated in this study, they can also be used with small animal imaging platforms to monitor the chondrogenic activity of implanted progenitors within orthotopic models of bone and cartilage repair.

Background: Several reports of severe infections associated with allograft tissue in knee reconstructive surgery have led many surgeons to consider routine intraoperative culture of allograft tissue before implantation. Thus, the purpose of this study was to determine the prevalence of positive soft tissue allograft cultures in reconstructive knee surgery, and evaluate its association with surgical site infection., Methods: Retrospective study of 202 patients who underwent knee reconstructive ligament surgeries, including revisions, between January 2013 and July 2017. Intraoperative culture results were obtained and the report of a surgical site infection during follow-up was recorded. Patients without cultures were excluded. A priori power analysis was performed. The association between positive culture results and development of surgical site infection was evaluated using Fisher's Exact test (P < 0.05)., Results: A total of 300 allografts were implanted in 202 patients. Mean average follow-up was 32.9 ± 12.5 (range 13 to 57.9) months. Sixteen patients had positive intraoperative allograft cultures (7.9%). The most frequently isolated organism was Bacillus species (six cultures); none of these patients presented with clinical signs of infection. Nine patients developed surgical site infections and were treated with oral antibiotics, and one patient developed septic arthritis that required surgical debridement of the implanted graft; all of these patients had a negative soft tissue allograft culture. No significant association was found between a positive culture and surgical site infection (P = 0.43)., Conclusion: There was no apparent association between positive intraoperative irradiated soft tissue allograft cultures and surgical site infection in reconstructive knee surgery., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Disease-modifying osteoarthritis drugs (DMOADs) should reach their intra-tissue target sites at optimal doses for clinical efficacy. The dense, negatively charged matrix of cartilage poses a major hindrance to the transport of potential therapeutics. In this work, electrostatic interactions were utilised to overcome this challenge and enable higher uptake, full-thickness penetration and enhanced retention of dexamethasone (Dex) inside rabbit cartilage. This was accomplished by using the positively charged glycoprotein avidin as nanocarrier, conjugated to Dex by releasable linkers. Therapeutic effects of a single intra-articular injection of low dose avidin-Dex (0.5 mg Dex) were evaluated in rabbits 3 weeks after anterior cruciate ligament transection (ACLT). Immunostaining confirmed that avidin penetrated the full cartilage thickness and was retained for at least 3 weeks. Avidin-Dex suppressed injury-induced joint swelling and catabolic gene expression to a greater extent than free Dex. It also significantly improved the histological score of cell infiltration and morphogenesis within the periarticular synovium. Micro-computed tomography confirmed the reduced incidence and volume of osteophytes following avidin-Dex treatment. However, neither treatment restored the loss of cartilage stiffness following ACLT, suggesting the need for a combinational therapy with a pro-anabolic factor for enhancing matrix biosynthesis. The avidin dose used caused significant glycosaminoglycan (GAG) loss, suggesting the use of higher Dex : avidin ratios in future formulations, such that the delivered avidin dose could be much less than that shown to affect GAGs. This charge-based delivery system converted cartilage into a drug depot that could also be employed for delivery to nearby synovium, menisci and ligaments, enabling clinical translation of a variety of DMOADs.

For evaluation of new approaches to drug delivery into cartilage, the choice of an animal model is critically important. Since cartilage thickness varies with animal size, different levels of drug uptake, transport and retention should be expected. Simple intra-articular injection can require very high drug doses to achieve a concentration gradient high enough for drug diffusion into cartilage. New approaches involve nanoparticle delivery of functionalized drugs directly into cartilage; however, diffusion-binding kinetics proceeds as the square of cartilage thickness. In this study, we demonstrate the necessity of using larger animals for sustained intra-cartilage delivery and retention, exemplified by intra-articular injection of Avidin (drug-carrier) into rabbits and compared to rats in vivo. Penetration and retention of Avidin within cartilage is greatly enhanced by electrostatic interactions. Medial tibial cartilage was the thickest of rabbit cartilages, which generated the longest intra-cartilage half-life of Avidin (τ1/2 = 154 h). In contrast, Avidin half-life in thinner rat cartilage was 5-6 times shorter (τ1/2 ∼ 29 h). While a weak correlation (R(2) = 0.43) was found between Avidin half-lives and rabbit tissue GAG concentrations, this correlation improved dramatically (R(2) = 0.96) when normalized to the square of cartilage thickness, consistent with the importance of cartilage thickness to evaluation of drug delivery and retention., (© 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Intra-articular (i.a.) drug delivery for local treatment of osteoarthritis remains inadequate due to rapid clearance by the vasculature or lymphatics. Local therapy targeting articular cartilage is further complicated by its dense meshwork of collagen and negatively charged proteoglycans, which can prevent even nano-sized solutes from entering. In a previous in vitro study, we showed that Avidin, due to its size (7 nm diameter) and high positive charge (pI 10.5), penetrated the full thickness of bovine cartilage and was retained for 15 days. With the goal of using Avidin as a nano-carrier for cartilage drug delivery, we investigated its transport properties within rat knee joints. Avidin penetrated the full thickness of articular cartilage within 6 h, with a half-life of 29 h, and stayed inside the joint for 7 days after i.a. injection. The highest concentration of Avidin was found in cartilage, the least in patellar tendon and none in the femoral bone; in contrast, negligible Neutravidin (neutral counterpart of Avidin) was present in cartilage after 24 h. A positive correlation between tissue sGAG content and Avidin uptake (R(2)  = 0.83) confirmed the effects of electrostatic interactions. Avidin doses up to at least 1 µM did not affect bovine cartilage explant cell viability, matrix catabolism or biosynthesis., (© 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Purpose: The objective of this study was to determine whether using mesenchymal stem cells (MSC) seeded in a collagen type I scaffold would be sufficient to regenerate the torn anterior cruciate ligament (ACL)., Methods: Anterior cruciate ligament transection was performed on both knees in 10 New Zealand rabbits and then repaired with as follows: suture alone (suture-treated group, n = 6), suture associated with collagen type I scaffold (collagen type I scaffold-treated group, n = 8) or suture associated with autologous MSC seeded on collagen type I scaffold (MSC/collagen type I scaffold-treated group, n = 6). At 12-week post-intervention, the animals were killed and the ACLs were characterised macroscopically and histologically. Data of the 3 groups were against normal ACL (normal group, n = 10)., Results: Macroscopic observation found that in MSC/collagen type I scaffold group, 33% of specimens showed a complete ACL regeneration, with a tissue similar to the normal ACL. Regeneration was not observed in the group treated with suture alone or associated with collagen type I scaffold without cells. In the latter, only a reparative attempt at the ends was observed. Histological analysis of the regenerated ACL showed a tissue with organised collagen and peripheric vessels., Conclusions: These results provide evidence that the use of MSC seeded in a collagen type I scaffold in the treatment of ACL injuries is associated with an enhancement of ligament regeneration. This MSC-based technique is a potentially attractive tool for improving the treatment of ACL ruptures.

Background: Mitral valve repair is considered better than mitral valve replacement for degenerative mitral regurgitation., Aim: To evaluate late clinical results of mitral valve repair as compared to mitral valve replacement in patients with degenerative mitral regurgitation., Patients and Methods: All patients subjected to open heart surgery for degenerative mitral regurgitation between 1990 and 2002 were assessed for surgical mortality, late cardiac and overall mortality, reoperation, readmission to hospital, functional capacity and anticoagulant therapy. Eighty eight patients (48 males) had mitral valve repair and 28 (19 males) had mitral valve replacement (23 with a mechanical prosthesis). Mean age was 59.9 +/- 14.8 (SD) and 61.3 +/- 14.6 years, respectively. Sixty three percent of patients with repair and 50% of those with valve replacement were in functional class III or IV before surgery., Results: Operative mortality was 2.3% for mitral valve repair and 3.6% for mitral valve replacement (NS). Also, there was no statistical difference in the need of reoperation during the follow-up period between both procedures (2.3% and 0%, respectively). Ninety four percent of the replacement patients but only 26% of the repair patients were in anticoagulant therapy at the end of the follow-up period (p < 0.001). Ten years survival rates were 82 +/- 6% for mitral valve repair and 54 +/- 11% for replacement. The corresponding cardiac related survival rates were 89 +/- 6% and 79 +/- 10%. At the end of follow-up, all surviving patients were in functional class I or II. Ten years freedom from cardiac event rates (death, cardiac related rehospitalization and reoperation) were 90 +/- 3% for mitral valve repair and 84 +/- 6% for replacement., Conclusion: Repair of the mitral valve offers a better overall survival and a better chance of freedom from cardiac events as well as need for anticoagulation 10 years after surgery.

Background: Surgical valve repair is a good alternative for correction of incompetent bicuspid aortic valve., Aim: To report the early and late surgical, clinical and ecochardiographic results of surgical repair of incompetent bicuspid aortic valves., Patients and Methods: Retrospective review of medical records of 18 patients aged 19 to 61 years, with incompetent bicuspid aortic valve in whom a valve repair was performed. Four patients had infectious endocarditis and 17 were in functional class I or II. Follow up ranged from 3 to 113 months after surgery., Results: A triangular resection of the prolapsing larger cusp, which included the middle raphe, was performed in 17 cases; in 13 of these, a complementary subcommisural annuloplasty was performed. In the remaining case, with a perforation of the non-coronary cusp, a pericardial patch was implanted; this procedure was also performed in 2 other cases. In 3 cases large vegetations were removed. Postoperative transesophageal echocardiography showed no regurgitation in 11 patients (62%) and mild regurgitation in 7 (38%). There was no operative morbidity or mortality. There were no deaths during the follow-up period. In 3 patients (17%) the aortic valve was replaced with a mechanical prosthesis, 8 to 108 months after the first operation. Reoperation was not needed in 93%+/-6,4% at 1 year and 85%+/-9,5% at 5 years, these patients were all in functional class I at the end of the follow-up period. 60% had no aortic regurgitation, 20% had mild and 20% moderate aortic regurgitation on echocardiographic examination. A significant reduction of the diastolic diameter of the left ventricle was observed, but there were no significant changes in systolic diameter or shortening fraction., Conclusions: Surgical repair of incompetent bicuspid aortic valves has low operative morbidity and mortality and has a low risk of reoperation.

Background and Objective: Sclerostomy ab externo with pulsed laser systems is currently in phase II clinical trials. The authors investigated the ablation dynamics of tissue treated with pulsed laser systems in the mid-infrared range to estimate the extent of thermo-mechanical damage to the sclera and the anterior chamber., Materials and Methods: Freshly harvested porcine eyes were used. A bare 400-micron fiber in direct contact with tissue was used for fistulization. Polarization light microscopy, fast-flash photography, as well as optical and acoustic transients were performed for analysis., Results: Substantial mechanical tissue deformation and dissections were found during pulsed laser ablation. The mechanical damage range within tissue far exceeds the pure thermal damage zone. Aspheric cavitation bubbles of up to 3 mm in length penetrate the anterior chamber after perforation. The cavitation demonstrates a significantly larger time constant in tissue than in water., Conclusions: Early fistula occlusions due to iris adherences may be attributed to iris trauma caused by cavitation. In response to the findings of this study, the authors propose an automatic feedback system to control the ablation process and minimize secondary ocular tissue effects. With respect to the overall damage zones, a new continuous-wave, mid-infrared diode laser system seems to be superior to pulsed laser systems.

ACE-inhibitors are known to have special renal effects, i.e. they increase ERPF, decrease the filtration fraction and lower proteinuria. These effects can be due to a decrease in angiotensin II (AII) levels as well as an increase in bradykinin. New and more specific AII-receptor antagonists may help to distinguish between effects exerted by angiotensin II and those exerted by bradykinin. We investigated the effects of losartan in 9 patients with essential hypertension (sitting mean diastolic blood pressure 95-120 mmHg). Renal hemodynamics were measured by continuous inulin-and PAH-clearance (GFR and RPF) after stopping antihypertensive therapy for 1 week, followed by a 2-week placebo period and after a 4-week treatment phase with losartan (50 mg/die) followed by a therapy with an ACE-inhibitor (ramipril 5mg/die). Additionally, urine albumin excretion (UAE) was measured. Treatment of patients with essential hypertension with losartan resulted in a significant decrease of MAP after three weeks of treatment (121 +/- 8 mmHg under placebo and 114 +/- 10 mmHg under losartan; * = p < 0.05). MAP after four weeks of losartan treatment was 115 +/- 11 mmHg. Regarding changes in renal hemodynamics we could not demonstrate a significant change for neither losartan nor the ACE-inhibitor. Urine albumin excretion was reduced by both treatment regimens in correlation to the magnitude of blood pressure reduction. Our data indicate that losartan induced a significant reduction in MAP in patients with essential arterial hypertension with only moderate effects on renal hemodynamics.

Patients with end-stage renal failure frequently develop venous stenoses or occlusions in their haemodialysis access fistulas caused by intimal fibrosis. A complete dilation with high pressure balloons up to 20 atm may be unsuccessful in such cases. We investigated two pulsed dye laser devices for the ablation of obstructions, which were not adequately treatable with a previous balloon angioplasty. From 11/90 to 4/92 a total amount of 154 PTAs of haemodialysis access fistulas were performed. In 23 of them additive laser angioplasty was necessary. 20 patients with Cimino fistulas presented 28 stenoses and two occlusions, whereas all the three patients with Goretex loops presented with occlusions. Two pulsed dye laser devices emitted at the wavelengths 504 nm (green) and 595 nm (red). Technical success was achieved in 22/23 cases, but clinical success was obtained in only 20/23 patients, due to two early reocclusions caused by thrombosis. 5 restenoses occurred two, three, 10, and twice 14 months after angioplasty with a mean follow-up period of 13.5 (5-18) months. Pulsed dye laser ablation in haemodialysis access fistula lesions due to intimal fibrosis greatly enriches radiological recanalisation techniques and is a valuable alternative to surgery if stand-alone balloon PTA fails.

A fistula from the anterior chamber of the eye into the subconjunctival space can be created by external laser application (laser sclerostomy). A pulsed (200 microseconds) Erbium-YAG laser was used because of the high scleral absorption. The laser energy was applied via a special fiber, which was inserted into a special application cannula to guide it into the subconjunctival space. A total of 24 rabbit eyes were treated and examined histologically after various postoperative periods. In all cases a functioning fistula and a marked reduction of the IOP (mean value 13 mmHg to 2 mmHg) were detected. On average, 60 mJ was required. The zone of thermal damage was limited to 40 microns. Cornea and lens showed no evidence of damage after 1 day or after 4 weeks. In the rabbit model, the fistulas were closed within 2 weeks, and long-term persistence of the fistula must therefore be verified in a clinical study. The method presented is minimally invasive, requires only a short operation time, causes minimal trauma to the conjunctiva, and may become an alternative to traditional fistulization operations.

An ab externo fistulizing procedure (sclerostomy) was performed in vivo in 48 rabbit eyes using a pulsed (20 microseconds and 200 microseconds) Holmium:YAG (Ho:YAG) laser (2120 nm) and a pulsed (200 microseconds) Erbium:YAG (Er:YAG) laser (2940 nm). The laser energy was delivered via an infrared transmitting fiber in contact with the sclera, with the fiber inserted into a specially sharpened retractable cannula in order to guide it into the subconjunctival space. An additional optical fiber-fiber coupling system was required for the Er-YAG laser. A patent fistula (200-micrometers diameter) with formation of a filtering bleb and marked intraocular pressure reduction was achieved in nearly all cases. Significant intraoperative or postoperative complications occurred only with the 20-microsecond Ho:YAG laser. Trauma to the conjunctiva was as minimal as in a subconjunctival injection. The Er:YAG laser created a smaller thermal damage zone at the fistula walls and required significantly less laser energy than the Ho-YAG laser.