High-throughput analysis of CircRNA in cows with naturally infected Staphylococcus aureus mammary gland.

Circular RNAs (CircRNA) are a special type of non-coding RNA molecule with a closed ring structure and are not affected by RNA exonucases. It has stable expression, is not easy to degrade, and exists in most eukaryotes. However, circRNA regulation of cow mastitis has not been widely recognized. Mammary epithelial tissues were collected from healthy Holstein cows (HCN) and mastitis Holstein cows (HCU). RNA sequencing (RNA SEQ) was performed for the differentially expressed circRNAs, and analysis results showed that 19 differentially expressed circRNAs were identified in HCN and HCU, among which 6 circRNAs were up-regulated and 13 circRNAs were down-regulated. We randomly selected nine circRNAs for Q-PCR verification, and the results showed consistent expression. Three circRNAs: circRNA2860, circRNA5323 and circRNA4027 were confirmed to be significantly differentially expressed circRNAs in cow mastitis. Also, their host genes TRPS1, SLC12A2 and MYH11 might be directly or indirectly play a role in cow mastitis. Furthermore, RNA polymerase transcription factor binding and tight junction are most enriched in GO and KEGG pathways, respectively. In addition, the regulatory network of circRNA-miRNA has been inferred from a bioinformatics perspective, which may help to understand the underlying molecular mechanism of circRNAs involved in regulating mastitis in cows.

Intracardiac amorphous tumor presenting in a patient with homocystinuria; a case report with literature review.

Homocystinuria is a rare genetic disease with autosomal recessive pattern. It is reported to be highest in Arabian descend and could cause thrombosis, but mainly peripherally. Cardiac amorphous tumor has been recognized in the past 20 years and it is also a very rare cause primary benign tumor of the heart. Most of the cases reported to be associated with end-stage renal disease. Homocystinuria associated with Cardiac Amorphous tumor is extremely rare. Up to our knowledge, there has been only one other case has been reported. Our patient is a 14-year-old female known case of homocystinuria presented with dyspnea and leg edema. On workup was found to have a mass in the right atrium extending to superior vena cava and inferior cava. Surgery undertaken on cardiopulmonary bypass partial resection of the mass was done and result came back as cardiac amorphous tumor. We assume the cause of this sinister complication of her primary illness is calcification of thrombus as stated in literature. And also recommend further studies regarding issue on hand.

Extra-Hepatic Hepatoid Carcinomas in Female Reproductive System: Three Case-Reports with a Literature Review.

PURPOSE: Hepatoid carcinoma of the ovary (HCO) and hepatoid carcinoma of the uterus (HCU) are rare malignancies that can be difficult to distinguish from other diseases such as hepatocellular carcinoma. In extremely rare cases, patients are negative for α-fetoprotein (AFP) by immunohistochemistry. Here we report 3 cases of HC of the female reproductive system, including 1 that was negative for AFP. PATIENTS AND METHODS: Three women aged 48, 56, and 67 years were treated at Qilu Hospital of Shandong University for HCO or HCU. We describe these cases in detail, including clinical features, diagnosis, treatment, and outcome, and review similar cases reported in the literature. RESULTS: All of our patients underwent surgery including hysterectomy and bilateral adnexectomy, and were treated with platinum-based chemotherapy. One patient died 3 months after the operation, and the other 2 are alive 22 and 63 months post surgery. CONCLUSION: The first-choice treatment for HCO and HCU is staging surgery, which should be followed by platinum-based chemotherapy.

Early Development of Newborn Screening for HCU and Current Challenges.

Classic homocystinuria (HCU) was added to newborn screening (NBS) by Robert Guthrie a few years after the disorder was first described. The justification for NBS was similar to that for PKU, that presymptomatic identification and early dietary treatment would prevent the clinical consequences, which, for HCU, are mental deficiency, ectopia lentis, skeletal abnormalities, and thromboembolism. It was assumed that identifying increased methionine in the screening blood specimen would identify all affected neonates. However, it is now clear that many with HCU are missed by NBS, mainly because the methionine level in the first days of life is normal or below the cutoff level in the NBS program. This includes virtually all of those with B6-responsive HCU. Thus, a more effective method of NBS for HCU should be considered. Included among the possibilities are decreasing the methionine cutoff level, requiring an increase in the Met/Phe ratio if the methionine level is not at or greater than the cutoff level, using methionine as the primary screen with homocysteine as a second-tier test, or replacing methionine with homocysteine as the primary screen. Homocysteine is the primary metabolite that increases in HCU, while the methionine increase is secondary, so homocysteine is usually increased before the increase in methionine, almost always during the first few days of life. Finally, targeted gene screening might be considered. All of these possibilities would impose added expense and labor to NBS, so meeting these challenges would likely require a regional or national effort.

Diagnosis of Classic Homocystinuria in Two Boys Presenting with Acute Cerebral Venous Thrombosis and Neurologic Dysfunction after Normal Newborn Screening.

Homocystinuria, caused by cystathionine ß-synthase deficiency, is a rare inherited disorder involving metabolism of methionine. Impaired synthesis of cystathionine leads to accumulation of homocysteine that affects several organ systems leading to abnormalities in the skeletal, cardiovascular, ophthalmic and central nervous systems. We report a 14-month-old and a 7-year-old boy who presented with neurologic dysfunction and were found to have cerebral venous sinus thromboses on brain magnetic resonance imaging (MRI)/magnetic resonance venogram (MRV) and metabolic and hypercoagulable work-up were consistent with classic homocystinuria. The 14-month-old boy had normal newborn screening. The 7-year-old boy initially had an abnormal newborn screen for homocystinuria but second tier test that consisted of total homocysteine was normal, so his newborn screen was reported as normal. With the advent of expanded newborn screening many treatable metabolic disorders are detected before affected infants and children become symptomatic. Methionine is the primary target in newborn screening for homocystinuria and total homocysteine is a secondary target. Screening is usually performed after 24-48 h of life in most states in the US and some states perform a second screen as a policy on all tested newborns or based on when the initial newborn screen was performed. This is done in hopes of detecting infants who may have been missed on their first screen. In the United Kingdom, NBS using dried blood spot is performed at 5 to 8 days after birth. It is universally known that methionine is a poor target and newborn screening laboratories have used different cutoffs for a positive screen. Reducing the methionine cutoff increases the sensitivity but not necessarily the specificity of the test and increasing the cutoff will miss babies who may have HCU whose levels may not be high enough to be detected at their age of ascertainment. It is not clear whether adjusting methionine level to decrease the false negative rates combined with total homocysteine as a second-tier test can be used effectively and feasibly to detect newborns with HCU. Between December 2005 and December 2020, 827,083 newborns were screened in Kentucky by MS/MS. Kentucky NBS program uses the postanalytical tools offered by the Collaborative Laboratory Integrated Reports (CLIR) project which considers gestational age and birthweight. One case of classical homocystinuria was detected and two were missed on first and second tier tests respectively. The newborn that had confirmed classical homocystinuria was one of twenty-three newborns that were referred for second tier test because of elevated methionine (cutoff is >60 µmol/L) and/or Met/Phe ratio (cutoff is >1.0); all 23 dried blood spots had elevated total homocysteine. One of the subjects of this case report had a normal methionine on initial screen and the other had a normal second-tier total homocysteine level. The performance of methionine and total homocysteine as screening analytes for homocystinuria suggest that it may be time for newborn screening programs to consider adopting next generation sequencing (NGS) platforms as alternate modality of metabolic newborn screening. Because of cost considerations, newborn screening programs may not want to adopt NGS, but the downstream healthcare cost incurred due to missed cases and the associated morbidity of affected persons far exceed costs to newborn screen programs. Since NGS is becoming more widely available and inexpensive, it may be feasible to change testing algorithms to use Newborn Metabolic NGS as the primary mode of testing on dry blood specimens with confirmation with biochemical testing. Some commercial laboratories have Newborn Screening Metabolic gene panel that includes all metabolic disorders on the most comprehensive newborn screening panel in addition to many other conditions that are not on the panel. A more targeted NGS panel can be designed that may not cost much and eventually help avoid the pitfalls associated with delayed diagnosis and cost of screening.

Disseminated Mycobacterium chimaera Following Open-Heart Surgery, the Heater-Cooler Unit Worldwide Outbreak: Case Report and Minireview.

Invasive cardiovascular infections by Mycobacterium chimaera associated with open-heart surgery have been reported worldwide since 2013. Here, we report a case of a 61 year old man, without any other particular medical background, who underwent cardiac surgery for replacing part of the ascending aorta by a bio-prosthetic graft. Eighteen months later, the patient was painful at the lower back with fever. A pyogenic vertebral osteomyelitis due to M. chimaera associated to graft infection was diagnosed after 6 months of sub-acute infection. The patient presented a disseminated disease with cerebral lesions, chorioretinitis, and chronic renal failure. Despite adequate antimicrobial treatment and graft explantation, the patient died after 6 years. We reviewed the literature on M. chimaera infections associated with open-heart surgery. The worldwide outbreak has been explained by airborne bioaerosol generated by the 3T heater-cooler unit (HCU) used during cardiac by-pass surgical procedures. These infections are difficult to diagnose because of a long latency period (up to several years), with no specific symptoms and a highly specialized microbiological diagnosis. The treatment is based on antibiotics and surgery. These infections are also difficult to treat, since the mortality rate is high around 50%. Prevention is necessary by modifying the use of HCUs in operating rooms.

[Palliative care for children in onco-hematology: Role of a specific home-care team]. / Soins palliatifs pédiatriques et fin de vie d'enfants suivis en onco-hématologie : place de l'hospitalisation à domicile.

BACKGROUND/OBJECTIVE: Our home-care unit (HCU) is specialized for pediatric cancer patients and has a strong palliative care activity. We believe that the introduction of home-care services can influence the place of palliative care and of death as well as the length of hospitalization. We aimed at describing characteristics and care course of patients treated in our HCU, and tried to identify some factors contributing to home care at the end of life. DESIGN/METHODS: We conducted a retrospective, observational, monocentric study about patients in pediatric onco-hematology, treated at least one day in our home-care unit, who died between July 1st 2013 and December 31st 2015. Statistical analysis was descriptive and analytic. RESULTS: A total of 74 patients known by our HCU died during study period. Eight were excluded. Forty-three out of 66 patients died at home. During the last 3 months of life, oncology patients have significantly less classical hospitalization, when compared to hematology patients. The implication of general physicians (GP) and nurses and information given to the family increase the possibility for home death. No significant association was found between ages at death, distance between home and hospital, other life conditions and place of death. CONCLUSIONS: Our HCU has a strong palliative care activity and a high rate of children dying at home. Good collaborations between our pediatric onco-hematology team and our HCU as well as between our HCU and caregivers optimize palliative care.

Diversity in the incidence and spectrum of organic acidemias, fatty acid oxidation disorders, and amino acid disorders in Asian countries: Selective screening vs. expanded newborn screening.

BACKGROUND: Expanded newborn screening (ENBS) utilizing tandem mass spectrometry (MS/MS) for inborn metabolic diseases (IMDs), such as organic acidemias (OAs), fatty acid oxidation disorders, (FAODs), and amino acid disorders (AAs), is increasingly popular but has not yet been introduced in many Asian countries. This study aimed to determine the incidence rates of OAs, FAODs, and AAs in Asian countries and Germany using selective screening and ENBS records. MATERIALS AND METHODS: Selective screening for IMDs using gas chromatography-mass spectrometry and MS/MS was performed among patients suspected to be afflicted in Asian countries (including Japan, Vietnam, China, and India) between 2000 and 2015, and the results from different countries were compared. Similarly, ENBS results from Japan, South Korea, Taiwan, and Germany were compared. Additionally, the results of selective screening and ENBS in Japan were compared. RESULTS: Among 39,270 patients who underwent selective screening, IMDs were detected in 1170. Methylmalonic acidemia was frequently identified in several countries, including Japan (81/377 diagnosed IMDs), China (94/216 IMDs), and India (72/293 IMDs). In Vietnam, however, ß-ketothiolase deficiency was particularly frequent (33/250 IMDs). ENBS yielded differences in overall IMD rates by country: 1:8557 in Japan, 1:7030 in Taiwan, 1:13,205 in South Korea, and 1:2200 in Germany. Frequently discovered diseases included propionic acidemia (PPA) and phenylketonuria (PKU) in Japan, 3-methylcrotonyl-CoA carboxylase deficiency (MCCD) and PKU in Taiwan, MCCD and citrullinemia type I in South Korea, and PKU and medium-chain acyl-CoA dehydrogenase deficiency in Germany. Furthermore, in Japan, selective screening and ENBS yielded respective PPA frequencies of 14.7% and 49.4% among all organic acidemias. CONCLUSION: The incidence rates of IMDs vary by country. Moreover, the disease spectra of IMDs detected via selective screening differ from those detected via ENBS.

Serum sample containing endogenous antibodies interfering with multiple hormone immunoassays. Laboratory strategies to detect interference.

OBJECTIVES: Endogenous antibodies (EA) may interfere with immunoassays, causing erroneous results for hormone analyses. As (in most cases) this interference arises from the assay format and most immunoassays, even from different manufacturers, are constructed in a similar way, it is possible for a single type of EA to interfere with different immunoassays. Here we describe the case of a patient whose serum sample contains EA that interfere several hormones tests. We also discuss the strategies deployed to detect interference. SUBJECTS AND METHODS: Over a period of four years, a 30-year-old man was subjected to a plethora of laboratory and imaging diagnostic procedures as a consequence of elevated hormone results, mainly of pituitary origin, which did not correlate with the overall clinical picture. RESULTS: Once analytical interference was suspected, the best laboratory approaches to investigate it were sample reanalysis on an alternative platform and sample incubation with antibody blocking tubes. Construction of an in-house 'nonsense' sandwich assay was also a valuable strategy to confirm interference. In contrast, serial sample dilutions were of no value in our case, while polyethylene glycol (PEG) precipitation gave inconclusive results, probably due to the use of inappropriate PEG concentrations for several of the tests assayed. CONCLUSIONS: Clinicians and laboratorians must be aware of the drawbacks of immunometric assays, and alert to the possibility of EA interference when results do not fit the clinical pattern.

Characterization of two pathogenic mutations in cystathionine beta-synthase: different intracellular locations for wild-type and mutant proteins.

Cystathionine ß-synthase (CBS) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the condensation of homocysteine with serine to generate cystathionine. Homocystinuria is an autosomal recessive disorder commonly caused by a deficiency of CBS activity. Here, we characterized a novel CBS mutation (c.260C>A (p.T87N)) and a previously reported variant (c.700G>A (p.D234N)) found in Venezuelan homocystinuric patients, one nonresponsive and one responsive to vitamin B6. Both mutant proteins were expressed in vitro in prokaryotic and eukaryotic cells, finding lower soluble expression in HEK-293 cells (19% T87N and 23% D234N) compared to wild-type CBS. Residual activities obtained for the mutant proteins were 3.5% T87N and 43% D234N. Gel exclusion chromatography demonstrated a tendency of the T87N mutant to aggregate while the distribution of the D234N mutant was similar to wild-type enzyme. Using immunofluorescence microscopy, an unexpected difference in intracellular localization was observed between the wild-type and mutant proteins. While the T87N mutant exhibited a punctate appearance, the wild-type protein was homogeneously distributed inside the cell. Interestingly, the D234N protein showed both distributions. This study demonstrates that the pathogenic CBS mutations generate unstable proteins that are unable (T87N) or partially unable (D234N) to assemble into a functional enzyme, implying that these mutations might be responsible for the homocystinuria phenotype.