Grandparental genotyping enhances exome variant interpretation

Trio exome sequencing is a powerful tool in the molecular investigation of monogenic disorders and provides an incremental diagnostic yield over proband‐only sequencing, mainly due to the rapid identification of de novo disease‐causing variants. However, heterozygous variants inherited from unaffected parents may be inadvertently dismissed, although multiple explanations are available for such scenarios including mosaicism in the parent, incomplete penetrance, imprinting, or skewed X‐inactivation. We report three probands, in which a pathogenic or likely pathogenic variant was identified upon exome sequencing, yet was inherited from an unaffected parent. Segregation of the variants (in NOTCH1, PHF6, and SOX10) in the grandparent generation revealed that the variant was de novo in each case. Additionally, one proband had skewed X‐inactivation. We discuss the possible genetic mechanism in each case, and urge caution in data interpretation of exome sequencing data. We illustrate the utility of expanding segregation studies to the grandparent generation and demonstrate the impact on exome interpretation strategies, by showing that objective genotype data can overcome subjective parental report of lack of symptoms.     

Oligozoospermia, asthenozoospermia, and sperm abnormalities in ex-addict to heroin, morphine, and hashish

A 40-year-old male had been addicted to heroin, morphine, hashish, and other narcotics for 12 years. At examination, 2 years after abstinence from drugs, his semen analysis revealed oligozoospermia, asthenozoospermia, and morphologically abnormal spermatozoa such as "round-headed" and "kinked"--sperm with neck abnormalities and immature forms. There was no evidence of other morphological abnormalities or of the presence of morphologically normal sperm. A possible correlation is discussed between the long-lasting drug addiction and morphological sperm abnormality, endocrinological function, karyotype, and immunological status.     

Targeted Delivery of Doxorubicin via Sterically Stabilized Immunoliposomes: Pharmacokinetics and Biodistribution in Tumor-bearing Mice

Purpose. To evaluate benefits in tumor localization, availability, and noncancerous organ distribution of doxorubicin (DOX) delivered via small (120 nm) sterically stabilized immunoliposomes targeted against a tumor-associated antigen in fibrosarcoma-bearing mice. Methods. DOX-loaded liposomes were prepared with (i) specific monoclonal IgG₃ antibody (32/2, D-SSIL-32/2); (ii) non-specific IgG₃ (D-SSIL-IgG); or (iii) no IgG (D-SSL) on their surface. Equal DOX amounts were injected intravenously via each type of liposome into BALB/c mice carrying experimental lung metastases of a polyoma virus-induced fibrosarcoma (A9 etc 220) expressing a polyoma virus-induced tumor-associated antigen (PAA) on their surface. Metastases occurred mainly in lung. Mice were treated at 3 stages of tumor development (micrometastases, medium-size metastases, and large, necrotic metastases). Performance evaluation was based on time-dependent quantification of DOX and DOX metabolites (DOX-M) in lung tumor, noncancerous organs, and plasma. Results. (i) DOX delivered via both SSIL retained the prolonged circulation time typical of DOX delivered via D-SSL. (ii) DOX accumulation in noncancerous organs was similar for all preparations. Low levels of DOX-M were obtained for all three preparations in all organs except liver, suggesting a similar processing, (iii) Preparations differed in behavior in lung tumor depending on tumor size and microanatomy. Only at the micrometastases stage were the specifically targeted D-SSIL-32/2 superior to D-SSL and D-SSIL-IgG, delivering 2–4 times more drug into the tumor, (iv) DOX-M level in all three tumor stages was in the following order: D-SSIL-32/2 >> D-SSL >> D-SSIL-IgG, suggesting that DOX delivered as D-SSIL-32/2 is most available to tumor cells. Conclusions. The advantage of specific targeting of sterically stabilized liposomes is expressed mainly in increasing availability of DOX to tumor cells in a way which is dependent on tumor microanatomy. The impact of this advantage to therapeutic efficacy remains to be determined.     

Perinatal lethal hypophosphatasia; Clinical,radiologic and morphologic findings

Clinical, radiographic and morphologic analysis of nineteen cases of perinatal (lethal) hypophosphatasia was performed. Three families each had two affected offspring. All of the patients had lethal short limb dwarfism with very soft calvaria. Other clinical findings included polyhydramnios, blue sclerae and spurs in the mid-portion of the forearms and lower legs. Considerable variability was found in the skeletal radiographs. In addition to the well known radiographic features such as generalized decrease in the size of ossified bones with some bones not ossified at all, other changes observed included: 1) marked variability in the amount of bone ossification; 2) variability between patients as to which bones were most severely affected; 3) unusually dense, round, flattened, butterfly shaped; and saggitally clefted vertebral bodies; 4) variability in femoral shape including chromosome like, campomelic like, and shortening with or without metaphyseal cupping or irregularities; 5) osteochondral projections (Bowdler spurs) of the midshaft of the fibula and ulna. Recognition of the marked clinical and radiographic variability in this autosomal recessive lethal skeletal dysplasia is important for accurate genetic counseling and prenatal diagnosis.     

Leukocyte Esterase Versus ICM 2018 Criteria in the Diagnosis of Periprosthetic Joint Infection

BackgroundA leukocyte esterase (LE) test is inexpensive and provides real-time information about patients suspected of periprosthetic joint infections (PJIs). The 2018 International Consensus Meeting (ICM) recommends it as a diagnostic tool with a 2+ cutoff. There is still a lack of data revealing LE utility versus the ICM 2018 criteria for PJI.MethodsThis is a retrospective study of patients who underwent revision total hip and total knee arthroplasty at a single institution between March 2009 and December 2019. All patients underwent joint aspiration before the arthrotomy, and the LE strip test was performed on aspirated joint fluid. PJI was defined using the 2018 ICM criteria.ResultsAs per the 2018 ICM criteria, 78 patients were diagnosed with chronic PJI and 181 were not infected. An LE test with a cutoff of $\ge$1+ had a sensitivity of 0.744, a specificity of 0.906, a positive predictive value of 0.773, an accuracy of 0.825 (95% confidence interval 0.772-0.878), and a negative predictive value of 0.891. The positive likelihood ratio (LR+) was 7.917. Using an LE cutoff of 2 + had a sensitivity of 0.513, a specificity of 1.000, and an accuracy of 0.756 (95% confidence interval—0.812).ConclusionLE is a rapid and inexpensive test which can be performed at the bedside. Its performance is valuable as per ICM criteria. Based on the findings of this study and the given cohort, we suggest using the cutoff of LE1+ (result = negative or trace) as a point of care test to exclude infection, whereas LE at 2 + threshold has near absolute specificity for the diagnosis.     

Uterine Remnants and Pelvic Pain in Females with Mayer-Rokitansky-Küster-Hauser Syndrome

ObjectiveTo assess the association between pelvic pain and uterine remnants and review the management of pelvic pain in females with Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome.DesignRetrospective cohort.SettingDepartment of Obstetrics and Gynecology at a tertiary referring medical center.PatientsForty-eight females with MRKH presenting from 1997 to 2011 with anatomy confirmed by magnetic resonance imaging (MRI).InterventionsNone.Main Outcome MeasurePrevalence Of uterine remnants and the association of uterine remnants with pelvic pain in females with MRKH.ResultsOf the 48 females with MRKH, 23 (48%) had uterine remnants and 22 (46%) had pelvic pain. Presence of endometrium was associated with pelvic pain (RR = 2.3; 95% CI = 1.2-4.7) in females with MRKH. Of the females with MKRH and pain, 9/22 had laparoscopy, with endometriosis seen in 5/9 of the uterine remnants at stages higher than are usually seen in teenagers (56%). Nine patients with pain and uterine remnants (8 with endometrium, 1 without) had laparoscopic removal of uterine remnants with resolution of pain.ConclusionsGiven the high prevalence of uterine remnants in females with MRKH, anatomic evaluation with MRI should be considered when assessing the etiology of pelvic pain. Presence of endometrium within uterine remnants, and subsequent endometriosis, in females with MRKH may be associated with pelvic pain necessitating surgical or medical management.     

Innovation and Integrity: Desiderata and Future Directions for Prevention and Intervention Science

This article summarizes essential implications of the papers within this special issue and discusses directions for future prevention and intervention research on conceptual issues, methodological and transfer-related challenges and opportunities. We identify a need to move from programs to principles in intervention research and encourage the implementation of research on potential mechanisms underlying intervention effectiveness. In addition, current methodological issues in intervention research are highlighted, including advancements in methodology and statistical procedures, extended outcome assessments, replication studies, and a thorough examination of potential biases. We further discuss transfer-related issues, for example the need for more research on the flexibility and adaptability of programs and intervention approaches as well as more general problems in knowledge translation reasoning the need for enhanced communication between practitioners, policy makers, and researchers. Finally, we briefly touch on the need to discuss the relation between single intervention programs, the mental health system, and changes of contextual conditions at the macro level.     

SVEP1 plays a crucial role in epidermal differentiation

SVEP1 is a recently identified multidomain cell adhesion protein, homologous to the mouse polydom protein, which has been shown to mediate cell‐cell adhesion in an integrin‐dependent manner in osteogenic cells. In this study, we characterized SVEP1 function in the epidermis. SVEP1 was found by qRT‐PCR to be ubiquitously expressed in human tissues, including the skin. Confocal microscopy revealed that SVEP1 is normally mostly expressed in the cytoplasm of basal and suprabasal epidermal cells. Downregulation of SVEP1 expression in primary keratinocytes resulted in decreased expression of major epidermal differentiation markers. Similarly, SVEP1 downregulation was associated with disturbed differentiation and marked epidermal acanthosis in three‐dimensional skin equivalents. In contrast, the dispase assay failed to demonstrate significant differences in adhesion between keratinocytes expressing normal vs low levels of SVEP1. Homozygous Svep1 knockout mice were embryonic lethal. Thus, to assess the importance of SVEP1 for normal skin homoeostasis in vivo, we downregulated SVEP1 in zebrafish embryos with a Svep1‐specific splice morpholino. Scanning electron microscopy revealed a rugged epidermis with perturbed microridge formation in the centre of the keratinocytes of morphant larvae. Transmission electron microscopy analysis demonstrated abnormal epidermal cell‐cell adhesion with disadhesion between cells in Svep1‐deficient morphant larvae compared to controls. In summary, our results indicate that SVEP1 plays a critical role during epidermal differentiation.