Plasma Glucosylsphingosine in GBA1 Mutation Carriers with and without Parkinson's Disease.

BACKGROUND: Biallelic mutations in the GBA1 gene encoding glucocerebrosidase cause Gaucher's disease, whereas heterozygous carriers are at risk for Parkinson's disease (PD). Glucosylsphingosine is a clinically meaningful biomarker of Gaucher's disease but could not be assayed previously in heterozygous GBA1 carriers. OBJECTIVE: The aim of this study was to assess plasma glucosylsphingosine levels in GBA1 N370S carriers with and without PD. METHODS: Glucosylsphingosine, glucosylceramide, and four other lipids were quantified in plasma from N370S heterozygotes with (n = 20) or without (n = 20) PD, healthy controls (n = 20), idiopathic PD (n = 20), and four N370S homozygotes (positive controls; Gaucher's/PD) using quantitative ultra-performance liquid chromatography tandem mass spectrometry. RESULTS: Plasma glucosylsphingosine was significantly higher in N370S heterozygotes compared with noncarriers, independent of disease status. As expected, Gaucher's/PD cases showed increases in both glucocerebrosidase substrates, glucosylsphingosine and glucosylceramide. CONCLUSIONS: Plasma glucosylsphingosine accumulation in N370S heterozygotes shown in this study opens up its future assessment as a clinically meaningful biomarker of GBA1-PD. © 2021 International Parkinson and Movement Disorder Society.

Higher Urine bis(Monoacylglycerol)Phosphate Levels in LRRK2 G2019S Mutation Carriers: Implications for Therapeutic Development.

BACKGROUND: LRRK2 mutations are a common cause of dominantly inherited PD. Previous studies showed decreases in urine levels of didocohexaenoyl (22:6) bis(monoacylglycerol)phosphate in LRRK2-knockout mice and in non-human primates treated with LRRK2 kinase inhibitors. We hypothesized that urine levels of bis(monoacylglycerol)phosphate isoforms will be higher in individuals with a PD-causing gain-of-kinase function mutation, LRRK2 G2019S. The objective of this study was to investigate alterations in urinary phospholipids as biomarkers of LRRK2 mutations and Parkinson's disease status/phenotypes. METHODS: Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to assess 54 bioactive phospholipids in urine from the LRRK2 Cohort Consortium (n = 80). To confirm and extend the findings, urine from an independent LRRK2 cohort from Columbia University Irving Medical Center (n = 116) was used. Both cohorts were composed of LRRK2 G2019S carriers and non-carriers with and without PD. RESULTS: In each cohort, 4 bis(monoacylglycerol)phosphate isoforms (di-18:1-bis[monoacylglycerol]phosphate, didocohexaenoyl [22:6] bis[monoacylglycerol] phosphate, 2,2'-di-22:6-bis[monoacylglycerol]phosphate, and 2,2'-di-18:1-bis[monoacylglycerol]phosphate) were significantly higher (2.5- to 4.3-fold) in G2019S carriers compared with non-carriers. Interestingly, 2,2'-di-18:1-bis(monoacylglycerol)phosphate levels were marginally higher in LRRK2 carriers with PD than in those without PD (P = 0.045). Moreover, increased 2,2' and total di-22:6-bis(monoacylglycerol)phosphate were associated with worse cognitive status assessed by the Montreal Cognitive Assessment (P = 0.0033 and 0.0144, respectively). CONCLUSIONS: The observed association of bis(monoacylglycerol)phosphate isoforms with LRRK2 G2019S mutation, PD status among G2019S carriers, and correlation with cognitive decline suggest the potential use of urinary bis(monoacylglycerol)phosphate isoforms as biomarkers for clinical trials of LRRK2-targeted therapies. © 2019 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

Clinical features, microbiological epidemiology and recommendations for management of cellulitis in extremity lymphedema.

BACKGROUND: This high volume, single center study investigated the prevalence, bacterial epidemiology, and responsiveness to antibiotic therapy of cellulitis in extremity lymphedema. METHODS: From 2003 to 2018, cellulitis events from a cohort of 420 patients with extremity lymphedema were reviewed. Demographics, lymphedema grading, symptoms, inflammatory markers, cultures and antibiotic therapy regimens were compiled from cellulitis episodes data. Univariate and multivariate analyses were performed for detailed analysis. RESULTS: A total of 131 separate episodes of cellulitis were recorded from 43 (81.1%) lower limb and 10 (19.9%) upper limb lymphedema patients. The prevalence and recurrence rates for cellulitis in lymphedema patients were 12.6% (53 of 420) and 56.6% (30 of 53), respectively. The most common findings were increased limb circumference (127 of 131; 96.9%) and abnormal C-reactive protein (CRP) level (86 of 113; 76.1%). Blood cultures were obtained in 79 (60.3%) incidents, with 9 (11.4%) returning positive. Streptococcus agalactiae was the most isolated bacterium (5 of 9; 55.5%). CONCLUSIONS: The cellulitis prevalence and recurrence rate in extremity lymphedema were 12.6%, and 56.6%, respectively. Strongest indicators of cellulitis were increased affected limb circumference and elevated CRP level. Empiric antibiotic therapy began with coverage for Steptococcus species before broadening to anti-Methicillin-resistant Staphylococcus aureus and anti-Gram negatives if needed for effective treatment of extremity lymphedema cellulitis.

Validation of a multiplexed LC-MS/MS clinical assay to quantify insulin-like growth factor-binding proteins in human serum and its application in a clinical study.

Circulating insulin-like growth factor-binding proteins (IGFBPs) continue to gain attention as biomarkers of drug activities on insulin like growth factor (IGF)/IGF receptor signaling pathways. A multiplexed LC-MS/MS method was validated for the absolute quantitation of IGFBPs in human serum. The method was used to measure screening concentrations of IGFBPs in spinal and bulbar muscular atrophy (SBMA) patients in a phase 2 clinical trial. Concentrations of IGFBP 1, 2, 3, and 5 were simultaneously determined based on representative signature peptides derived from an optimized trypsin digestion procedure. Signature peptide levels were absolutely quantitated using a sensitive/specific targeted LC-MS/MS method. Corresponding mass-shifted, stable isotope-labeled peptides were employed as internal standards. A true blank matrix for the quantitation of IGFBPs was not available since they are endogenous proteins in human serum. In this method, calibration standards/curves were prepared using authentic synthetic peptides spiked into a surrogate matrix. The surrogate matrix was generated from human serum treated in the same way as the study samples, but using iodoacetic acid instead of iodoacetamide as the alkylation reagent. This surrogate matrix approach allowed for the direct and sensitive/specific quantification of IGFBP 1, 2, 3, and 5 due to the lack of any endogenous background. Equivalent matrix effect and recovery of analytes was achieved for the authentic and surrogate matrices. The fully validated LC-MS/MS assay will allow further evaluation of the utility of IGFBP biomarkers in clinical trials.

A multiplexed UPLC-MS/MS assay for the simultaneous measurement of urinary safety biomarkers of drug-induced kidney injury and phospholipidosis.

Drug-induced kidney injury (DIKI) is a major concern in drug risk assessment given its clinical importance and the absence of a sensitive/specific method of diagnosis. Pharmaceutical regulatory agencies have qualified and issued letters of support for new biomarkers to better evaluate DIKI in nonclinical toxicity and clinical studies. Additional efforts have focused on drug-induced phospholipidosis (DIPL) and its potential link with collateral renal damage. The combined use of urinary biomarkers is an efficient way to evaluate renal safety in nonclinical and clinical studies. Eight FDA/EMA/PMDA qualified (or supported) urinary biomarkers, including kidney injury molecule-1 (KIM-1), ß2-microglobulin (B2M), clusterin (CLU), cystatin C (CysC), trefoil factor 3 (TFF3), neutrophil gelatinase-associated lipocalin (NGAL), osteopontin (OPN), and alpha-glutathione S-transferase (α-GST), were quantified by multiplex UPLC-MS/MS in a repeat dose study of gentamicin in rats. Rats administered gentamicin at 100 mg/kg/day for 2 weeks developed renal lesions detected by histopathology. Biomarkers of tubular damage (CLU, KIM-1, OPN) increased 9.8, 34.7, and 35.6-fold (relative to concurrent controls), respectively, after 2 weeks of dosing. Biomarkers of glomerular damage and/or impairment of tubular reabsorption (CysC, B2M) increased 11.7 and 22.6-fold. NGAL and α-GST increased <3-fold after 2 weeks of dosing. TFF3 was comparable to concurrent controls. The elevated biomarker concentrations met PSTC threshold criteria and were consistent with mechanisms of gentamicin nephrotoxicity. Increased urinary di-22:6-BMP indicated concomitant DIPL as confirmed by TEM. This work provides evidence supporting the combined use of the DIKI biomarker panel and di-22:6-BMP as a biomarker of DIPL in drug risk assessment.

The Correlation of Age and Patterns of Maxillofacial Bone Fractures and Severity of Associated Injuries Caused by Motorcycle Accidents.

BACKGROUND: Every year, there are significant numbers of motorcycle accident casualties in Taiwan. These accidents are the leading cause of maxillofacial trauma. Age should be an important factor of maxillofacial fracture patterns yet there is limited literature on the topic. Therefore, this study aims to evaluate the correlation of age with maxillofacial fracture in motorcycle accidents. METHODS: This is a retrospective descriptive analysis conducted over 2-year period at Linkou Chang Gung Memorial Hospital. We focused on the population of maxillofacial injury caused by motorcycle accidents. Data, including demographics, age, fracture patterns of facial bones, and other associated injuries, were collected. RESULTS: Among 881 admissions, there were 179 patients in the minor group, 644 patients in the adult group, and 58 patients in the geriatric group. With patterns of maxillofacial fracture, midface fracture was the most common type. The minor group had higher incidence of mandibular fracture. The geriatric group sustained more midface fracture. Associated injuries, such as severe head injuries and c-spine injury, were more likely to occur with the old age victims. The overall mortality rate was 3.1%. CONCLUSIONS: Our study presents the different trends of fracture patterns in different age groups, which is associated with different types of treatment required. We summarized all these data in the hope of providing further assistance to trauma doctor dealing with motorcycle accidents.

Di-22:6-bis(monoacylglycerol)phosphate: A clinical biomarker of drug-induced phospholipidosis for drug development and safety assessment.

The inability to routinely monitor drug-induced phospholipidosis (DIPL) presents a challenge in pharmaceutical drug development and in the clinic. Several nonclinical studies have shown di-docosahexaenoyl (22:6) bis(monoacylglycerol) phosphate (di-22:6-BMP) to be a reliable biomarker of tissue DIPL that can be monitored in the plasma/serum and urine. The aim of this study was to show the relevance of di-22:6-BMP as a DIPL biomarker for drug development and safety assessment in humans. DIPL shares many similarities with the inherited lysosomal storage disorder Niemann-Pick type C (NPC) disease. DIPL and NPC result in similar changes in lysosomal function and cholesterol status that lead to the accumulation of multi-lamellar bodies (myeloid bodies) in cells and tissues. To validate di-22:6-BMP as a biomarker of DIPL for clinical studies, NPC patients and healthy donors were classified by receiver operator curve analysis based on urinary di-22:6-BMP concentrations. By showing 96.7-specificity and 100-sensitivity to identify NPC disease, di-22:6-BMP can be used to assess DIPL in human studies. The mean concentration of di-22:6-BMP in the urine of NPC patients was 51.4-fold (p ≤ 0.05) above the healthy baseline range. Additionally, baseline levels of di-22:6-BMP were assessed in healthy non-medicated laboratory animals (rats, mice, dogs, and monkeys) and human subjects to define normal reference ranges for nonclinical/clinical studies. The baseline ranges of di-22:6-BMP in the plasma, serum, and urine of humans and laboratory animals were species dependent. The results of this study support the role of di-22:6-BMP as a biomarker of DIPL for pharmaceutical drug development and health care settings.

Reconstruction of a Large Composite Hemicraniofacial Defect with Bipedicled DIEP/TRAM Flaps.

Head and neck reconstructions are often challenging due to the mix of functional and aesthetic goals. It can be tricky when different tissue types are required to reconstruct each subunit. Craniofacial reconstructions require a large volume of tissue to cover defects that span across a large convex area. The latissimus dorsi muscle flap is a workhorse free flap used frequently by surgeons due to its ability to cover over a large surface area. However, there are unique situations when even the latissimus dorsi muscle is not enough to provide the bulk cover for the craniofacial defect. We present a complex case of a hemicraniofacial reconstruction after a hemifacial orbital exenteration and cranial resection of a large neglected basal cell carcinoma.

Memory-Based Prediction Deficits and Dorsolateral Prefrontal Dysfunction in Schizophrenia.

BACKGROUND: Theories suggest that people with schizophrenia (SZ) have problems generating predictions based on past experiences. The dorsolateral prefrontal cortex (DLPFC) and hippocampus participate in memory-based prediction. We used functional magnetic resonance imaging to investigate DLPFC and hippocampal function in healthy control (HC) subjects and people with SZ during memory-based prediction. METHODS: Prior to scanning, HC subjects (n = 54) and people with SZ (n = 31) learned 5-object sequences presented in fixed or random orders on each repetition. During scanning, participants made semantic decisions (e.g., "Can this object fit in a shoebox?") on a continuous stream of objects from fixed and random sequences. Sequence prediction was demonstrated by faster semantic decisions for objects in fixed versus random sequences because memory could be used to anticipate and more efficiently process semantic information about upcoming objects in fixed sequences. Representational similarity analyses were used to determine how each sequence type was represented in the posterior hippocampus and DLPFC. RESULTS: Sequence predictions were reduced in individuals with SZ relative to HC subjects. Representational similarity analyses revealed stronger memory-based predictions in the DLPFC of HC subjects than people with SZ, and DLPFC representations correlated with more successful predictions in HC subjects only. For the posterior hippocampus, voxel pattern similarity was increased for fixed versus random sequences in HC subjects only, but no significant between-group differences or correlations with prediction success were observed. CONCLUSIONS: Individuals with SZ are capable of learning temporal sequences; however, they are impaired using memory to predict upcoming events as efficiently as HC subjects. This deficit appears related to disrupted neural representation of sequence information in the DLPFC.

LRRK2 and GBA1 variant carriers have higher urinary bis(monacylglycerol) phosphate concentrations in PPMI cohorts.

We quantified concentrations of three isoforms of the endolysosomal lipid, bis(monoacylglycerol) phosphate (BMP) in the urine of deeply phenotyped cohorts in the Parkinson's Progression Markers Initiative: LRRK2 G2019S PD (N = 134) and non-manifesting carriers (NMC) (G2019S+ NMC; N = 182), LRRK2 R1441G PD (N = 15) and R1441G+ NMC (N = 15), GBA1 N409S PD (N = 76) and N409S+ NMC (N = 178), sporadic PD (sPD, N = 379) and healthy controls (HC) (N = 190). The effects of each mutation and disease status were analyzed using nonparametric methods. Longitudinal changes in BMP levels were analyzed using linear mixed models. At baseline, all LRRK2 carriers had 3-7× higher BMP levels compared to HC, irrespective of the disease status. GBA1 N409S carriers also showed significant, albeit smaller, elevation (~30-40%) in BMP levels compared to HC. In LRRK2 G2019S PD, urinary BMP levels remained stable over two years. Furthermore, baseline BMP levels did not predict disease progression as measured by striatal DaT imaging, MDS-UPDRS III Off, or MoCA in any of the cohorts. These data support the utility of BMP as a target modulation biomarker in therapeutic trials of genetic and sPD but not as a prognostic or disease progression biomarker.

Elevated urine BMP phospholipids in LRRK2 and VPS35 mutation carriers with and without Parkinson's disease.

Elevated urine bis(monoacylglycerol)phosphate (BMP) levels have been found in gain-of-kinase function LRRK2 G2019S mutation carriers. Here, we have expanded urine BMP analysis to other Parkinson's disease (PD) associated mutations and found them to be consistently elevated in carriers of LRRK2 G2019S and R1441G/C as well as VPS35 D620N mutations. Urine BMP levels are promising biomarkers for patient stratification and potentially target engagement in clinical trials of emerging targeted PD therapies.

CRISPR screens for lipid regulators reveal a role for ER-bound SNX13 in lysosomal cholesterol export.

We report here two genome-wide CRISPR screens performed to identify genes that, when knocked out, alter levels of lysosomal cholesterol or bis(monoacylglycero)phosphate. In addition, these screens were also performed under conditions of NPC1 inhibition to identify modifiers of NPC1 function in lysosomal cholesterol export. The screens confirm tight coregulation of cholesterol and bis(monoacylglycero)phosphate in cells and reveal an unexpected role for the ER-localized SNX13 protein as a negative regulator of lysosomal cholesterol export and contributor to ER-lysosome membrane contact sites. In the absence of NPC1 function, SNX13 knockdown redistributes lysosomal cholesterol and is accompanied by triacylglycerol-rich lipid droplet accumulation and increased lysosomal bis(monoacylglycero)phosphate. These experiments provide unexpected insight into the regulation of lysosomal lipids and modification of these processes by novel gene products.

Reconstruction of Pelvic Exenteration Defect with Free Anterolateral Thigh Flap: A Case Report.

The pedicled anterolateral thigh flap, with or without the vastus lateralis muscle, has been described for pelvic exenteration defect reconstruction. However, its use as a free flap for this type of defect is not routinely followed. To reconstruct an extensive pelvic defect in the presence of two ostomies, we describe a free anterolateral thigh flap with deep inferior epigastric pedicles as recipient vessels.

Musculoseptocutaneous Perforator of Anterolateral Thigh Flap: A Clinical Study.

BACKGROUND: The anterolateral thigh flap is one of the most useful workhorse flaps for microsurgical reconstruction. However, it can pose a great challenge to surgeons because of its anatomical variability. As the technology advances, not only septocutaneous or musculocutaneous courses of anterolateral thigh perforators but also a hybrid musculoseptocutaneous perforator pattern have been identified on computerized imaging and on cadaveric study. However, there is a paucity of clinical study in the literature. The aim of this investigation was to identify the features of this pattern. METHODS: All patients undergoing anterolateral thigh flap harvest between September of 2017 and May of 2018 performed by a single surgeon are included. Every pulsatile perforator was dissected to document its location on the thigh, emerging location (septum/muscle), size, course, and origin. RESULTS: Thirty-seven patients with 115 perforators were identified. Ten percent of perforators were septocutaneous, 37 percent were musculoseptocutaneous, and 52 percent were musculocutaneous. Forty-seven percent of perforators emerged on the septum between the rectus femoris and the vastus lateralis. Eighty-one percent of patients had one or more perforators in the "hot zone." Medium and large perforators were more frequently located in the proximal and hot zones. All perforators originated from the vascular tree of the lateral circumflex femoral artery, with 10 percent originating from the transverse branch, 28 percent originating from the oblique branch, and 62 percent originating from the descending branch. CONCLUSIONS: A high proportion of musculoseptocutaneous perforators were identified. The clinical relevance of this is to be very cautious on the skin paddle design while harvesting the flap.

A new approach to preexisting vertical midline abdominal scars with crossover DIEP flap breast reconstruction.

Breast reconstruction using a free transverse rectus abdominis myocutaneous flap or a deep inferior epigastric perforator (DIEP) flap is a challenge in patients with a vertical midline abdominal scar due to the poor perfusion of the lower abdominal skin ellipse across the midline. In such patients, only one half of the abdominal skin ellipse can be used with certainty, and this limits the amount of tissue available for reconstructing the breast. Two cases of breast reconstruction in patients with a lower midline abdominal scar are presented using the DIEP flap, in which the poor perfusion across the midline scar was overcome by a technique of crossover anastomoses between the two deep inferior epigastric pedicles. Reliable perfusion of the entire lower abdominal skin ellipse was achieved. This crossover anastomoses technique overcomes the poor perfusion imposed by the vertical midline abdominal scar and enables DIEP flap breast reconstruction to be offered to women with midline abdominal scars.

Elucidation of potential bortezomib response markers in mutliple myeloma patients.

Liquid chromatography coupled to mass spectrometry (LC/MS) was used to elucidate early biomarkers of bortezomib response in multiple myeloma patients. The change in serum myeloma M-protein level, maintained for a minimum of 6 weeks, is used as one of the main criteria to evaluate patient clinical response to therapy. The objective of this study was to identify biomarkers using LC/MS in order to predict patient response to bortezomib sooner and more accurately compared to serum M-protein levels. The plasma LC/MS biomolecular/biochemical profiles, comprised of thousands of endogenous small molecules, peptides and proteins, were determined for 10 multiple myeloma patients at predose and 24 h after initial dosing with bortezomib. The comparative analysis of the metabolic profiles of non-responders and partial responders provided an opportunity to investigate mechanisms related to disease progression and identify biomarkers related to drug response. The plasma levels of two potential efficacy response markers were significantly more abundant in the non-responsive patients compared to the responders at 24-h postdose. The potential response biomarkers, apolipoprotein C-I and apolipoprotein C-I', were identified by mass spectral analyses and confirmed by authentic protein standards based on MALDI-TOF MS/MS sequencing of proteolytic peptides.