The essential role of centrosomal NDE1 in human cerebral cortex neurogenesis.

We investigated three families whose offspring had extreme microcephaly at birth and profound mental retardation. Brain scans and postmortem data showed that affected individuals had brains less than 10% of expected size (≤10 standard deviation) and that in addition to a massive reduction in neuron production they displayed partially deficient cortical lamination (microlissencephaly). Other body systems were apparently unaffected and overall growth was normal. We found two distinct homozygous mutations of NDE1, c.83+1G>T (p.Ala29GlnfsX114) in a Turkish family and c.684_685del (p.Pro229TrpfsX85) in two families of Pakistani origin. Using patient cells, we found that c.83+1G>T led to the use of a novel splice site and to a frameshift after NDE1 exon 2. Transfection of tagged NDE1 constructs showed that the c.684_685del mutation resulted in a NDE1 that was unable to localize to the centrosome. By staining a patient-derived cell line that carried the c.83+1G>T mutation, we found that this endogeneously expressed mutated protein equally failed to localize to the centrosome. By examining human and mouse embryonic brains, we determined that NDE1 is highly expressed in neuroepithelial cells of the developing cerebral cortex, particularly at the centrosome. We show that NDE1 accumulates on the mitotic spindle of apical neural precursors in early neurogenesis. Thus, NDE1 deficiency causes both a severe failure of neurogenesis and a deficiency in cortical lamination. Our data further highlight the importance of the centrosome in multiple aspects of neurodevelopment.

Online vs. traditional learning: A comparative analysis of student's responses during COVID-19.

BACKGROUND: The coronavirus 2019 (COVID-19) pandemic had a major impact on the educational institutes globally and resulted in the transition from traditional educational systems to online electronic learning methods. OBJECTIVE: This study endeavored to address and compare the dental student's understanding regarding the Oral Biology course by assessing their knowledge and skills between the students enrolled during 2020 (online teaching) and 2021 (face-to-face teaching). This study also aimed to assess the perception of dental students regarding the virtual mode of teaching. METHODS: This quasi-experimental study evaluated students from two years, in which a retrospective group of dental students enrolled during the academic year 2020 were compared with a prospective experimental group enrolled during the academic year 2021. Knowledge and skills for both years were compared. This comprised of theoretical and practical component. Students also completed a questionnaire that assessed student's perception regarding online teaching. RESULTS: A total of 98 students of Bahria University Dental College participated. A comparison between grade scores of online and traditional learning groups reported statistically significant differences (p = 0.05*) with regards to short answer question types, viva or verbal questions (p = 0.016*). Strong correlations were observed via the multivariable analysis. All correlations were statistically significant at p < 0.01 level. Learning satisfaction was not satisfactorily observed by the online learning group. CONCLUSION: It can be concluded that despite the disruption that the educational sector had to face due to the COVID-19 pandemic, based on the differences in the subjective domain scores, students appeared to like on-campus teaching more than the online teaching, as most students found online learning to be stressful and were quite unsatisfied. However, the difference in mode of teaching did not affect the knowledge and skills of the dental students.

Previously described sequence variant in CDK5RAP2 gene in a Pakistani family with autosomal recessive primary microcephaly.

BACKGROUND: Autosomal Recessive Primary Microcephaly (MCPH) is a disorder of neurogenic mitosis. MCPH leads to reduced cerebral cortical volume and hence, reduced head circumference associated with mental retardation of variable degree. Genetic heterogeneity is well documented in patients with MCPH with six loci known, while pathogenic sequence variants in four respective genes have been identified so far. Mutations in CDK5RAP2 gene at MCPH3 locus have been least involved in causing MCPH phenotype. METHODS: All coding exons and exon/intron splice junctions of CDK5RAP2 gene were sequenced in affected and normal individuals of Pakistani MCPH family of Kashmiri origin, which showed linkage to MCPH3 locus on chromosome 9q33.2. RESULTS: A previously described nonsense mutation [243 T>A (S81X)] in exon 4 of CDK5RAP2 gene has been identified in the Pakistani family, presented here, with MCPH Phenotype. Genomic and cDNA sequence comparison revealed that the exact nomenclature for this mutation is 246 T>A (Y82X). CONCLUSION: Recurrent observation of Y82X mutation in CDK5RAP2 gene in this Pakistani family may be a sign of confinement of a rare ancestral haplotype carrying this pathogenic variant within Northern Pakistani population, as this has not been reported in any other population.

Sweeper's lung disease: a cross-sectional study of an overlooked illness among sweepers of Pakistan.

BACKGROUND: Sweepers are prone to develop chronic obstructive pulmonary disease even without tobacco smoking. PURPOSE: To investigate roadside dust as a cause of air flow obstruction among sweepers, and the role of spirometry in its preclinical diagnosis. MATERIAL AND METHODS: One-hundred nonsmoking sweepers (aged 30-60 years) of both sexes sweeping on roads for 8-12 hours a day for the Capital Development Authority of Islamabad, Pakistan were used as study participants (Group A). One-hundred healthy nonsmokers (aged 30-60 years) in the same socioeconomic group and living in the same environment represented the nonsweeper group (Group B). After proper clinical evaluation and chest X-rays, spirometric evaluation was carried out in both groups. Comparisons were drawn between various spirometric parameters. RESULTS: Pulmonary function tests showed that the mean forced vital capacity was 78 ± 1.40 in the sweeper group (Group A) and 83 ± 0.86 in the nonsweeper group (Group B). Mean forced expiratory volume in 1 second was 66 ± 1.67 in Group A and 85 ± 0.85 in Group B (P < 0.05), a difference of 19%. The forced midexpiratory flow was 41% lower in Group A than in Group B (P < 0.0001). The pattern of pulmonary function obstruction was shown to be proportional to the duration of exposure to dust caused by sweeping. CONCLUSION: Occupational exposure to dust leads to an obstructive pattern among sweepers. Spirometry is the simplest, noninvasive technique to detect preclinical disease.

A primary microcephaly protein complex forms a ring around parental centrioles.

Autosomal recessive primary microcephaly (MCPH) is characterized by a substantial reduction in prenatal human brain growth without alteration of the cerebral architecture and is caused by biallelic mutations in genes coding for a subset of centrosomal proteins. Although at least three of these proteins have been implicated in centrosome duplication, the nature of the centrosome dysfunction that underlies the neurodevelopmental defect in MCPH is unclear. Here we report a homozygous MCPH-causing mutation in human CEP63. CEP63 forms a complex with another MCPH protein, CEP152, a conserved centrosome duplication factor. Together, these two proteins are essential for maintaining normal centrosome numbers in cells. Using super-resolution microscopy, we found that CEP63 and CEP152 co-localize in a discrete ring around the proximal end of the parental centriole, a pattern specifically disrupted in CEP63-deficient cells derived from patients with MCPH. This work suggests that the CEP152-CEP63 ring-like structure ensures normal neurodevelopment and that its impairment particularly affects human cerebral cortex growth.

Mutation analysis of the ASPM gene in 18 Pakistani families with autosomal recessive primary microcephaly.

Autosomal recessive primary microcephaly (MCPH) is a rare neurological disorder, in which the patients exhibit reduced occipital frontal head circumference (>3 standard deviations) and mild-to-severe mental retardation. Autosomal recessive primary microcephaly is genetically heterogeneous and 7 loci have been reported to date. Mutations in ASPM (abnormal spindle-like, microcephaly associated) gene are the most common cause of autosomal recessive primary microcephaly in the majority of the reported families. In the current investigation, we have located and studied 21 families with autosomal recessive primary microcephaly. Genotyping using polymorphic microsatellite markers linked to 7 autosomal recessive primary microcephaly loci revealed linkage of 18 families to the MCPH5 locus. Sequence analysis of the ASPM gene in 18 linked families detected 2 novel nonsense mutations (c.2101C>T/p.Q701X; c.9492T>G/p.Y3164X) in 2 families and 2 novel deletion mutations (c.6686delGAAA/p.R2229TfsX9; c.77delG/p.G26AfsX41) in 2 other families. Three previously described mutations (c.3978G>A/p.W1326X; c.1260delTCAAGTC/p.S420SfsX32; c.9159delA/p.K3053NfsX4) were also detected in 11 families. These identified mutations extended the body of evidence implicating the ASPM gene in the pathogenesis of human hereditary primary microcephaly.

WDR62 is associated with the spindle pole and is mutated in human microcephaly.

Autosomal recessive primary microcephaly (MCPH) is a disorder of neurodevelopment resulting in a small brain. We identified WDR62 as the second most common cause of MCPH after finding homozygous missense and frame-shifting mutations in seven MCPH families. In human cell lines, we found that WDR62 is a spindle pole protein, as are ASPM and STIL, the MCPH7 and MCHP7 proteins. Mutant WDR62 proteins failed to localize to the mitotic spindle pole. In human and mouse embryonic brain, we found that WDR62 expression was restricted to neural precursors undergoing mitosis. These data lend support to the hypothesis that the exquisite control of the cleavage furrow orientation in mammalian neural precursor cell mitosis, controlled in great part by the centrosomes and spindle poles, is critical both in causing MCPH when perturbed and, when modulated, generating the evolutionarily enlarged human brain.