Homozygosity for a missense variant in COMP gene associated with severe pseudoachondroplasia.

The phenotypic spectrum associated with heterozygous mutations in cartilage oligomeric matrix protein gene (COMP) range from a mild form of multiple epiphyseal dysplasia (MED) to pseudoachondroplasia (PSACH). However, the phenotypic effect from biallelic COMP variants is unclear. We investigated a large consanguineous Pakistani family with a severe form of PSACH in 2 individuals. Another 14 family members presented with a mild PSACH phenotype similar to MED. Using exome sequencing and subsequent segregation analysis, we identified homozygosity for a COMP missense variant [c.1423G>A; p.(D475N)] in the 2 severely affected individuals, whereas family members with the mild PSACH phenotype were heterozygous. Our observations show for the first time that a biallelic COMP variant may be associated with pronounced and widespread skeletal malformations suggesting an additive effect of the 2 mutated alleles.

Genetic heterogeneity in Pakistani microcephaly families revisited.

Autosomal recessive primary microcephaly (MCPH) is a rare and heterogeneous genetic disorder characterized by reduced head circumference, low cognitive prowess and, in general, architecturally normal brains. As many as 14 different loci have already been mapped. We recruited 35 MCPH families in Pakistan and could identify the genetic cause of the disease in 31 of them. Using homozygosity mapping complemented with whole-exome, gene panel or Sanger sequencing, we identified 12 novel mutations in 3 known MCPH-associated genes - 9 in ASPM, 2 in MCPH1 and 1 in CDK5RAP2. The 2 MCPH1 mutations were homozygous microdeletions of 164,250 and 577,594 bp, respectively, for which we were able to map the exact breakpoints. We also identified four known mutations - three in ASPM and one in WDR62. The latter was initially deemed to be a missense mutation but we demonstrate here that it affects splicing. As to ASPM, as many as 17 out of 27 MCPH5 families that we ascertained in our sample were found to carry the previously reported founder mutation p.Trp1326*. This study adds to the mutational spectra of four known MCPH-associated genes and updates our knowledge about the genetic heterogeneity of MCPH in the Pakistani population considering its ethnic diversity.

Microduplications encompassing the Sonic hedgehog limb enhancer ZRS are associated with Haas-type polysyndactyly and Laurin-Sandrow syndrome.

Laurin-Sandrow syndrome (LSS) is a rare autosomal dominant disorder characterized by polysyndactyly of hands and/or feet, mirror image duplication of the feet, nasal defects, and loss of identity between fibula and tibia. The genetic basis of LSS is currently unknown. LSS shows phenotypic overlap with Haas-type polysyndactyly (HTS) regarding the digital phenotype. Here we report on five unrelated families with overlapping microduplications encompassing the Sonic hedgehog (SHH) limb enhancer ZPA regulatory sequence (ZRS) on chromosome 7q36. Clinically, the patients show polysyndactyly phenotypes and various types of lower limb malformations ranging from syndactyly to mirror image polydactyly with duplications of the fibulae. We show that larger duplications of the ZRS region (>80 kb) are associated with HTS, whereas smaller duplications (<80 kb) result in the LSS phenotype. On the basis of our data, the latter can be clearly distinguished from HTS by the presence of mirror image polysyndactyly of the feet with duplication of the fibula. Our results expand the clinical phenotype of the ZRS-associated syndromes and suggest that smaller duplications (<80 kb) are associated with a more severe phenotype. In addition, we show that these small microduplications within the ZRS region are the underlying genetic cause of Laurin-Sandrow syndrome.

Identifying human disease genes: advances in molecular genetics and computational approaches.

The human genome project is one of the significant achievements that have provided detailed insight into our genetic legacy. During the last two decades, biomedical investigations have gathered a considerable body of evidence by detecting more than 2000 disease genes. Despite the imperative advances in the genetic understanding of various diseases, the pathogenesis of many others remains obscure. With recent advances, the laborious methodologies used to identify DNA variations are replaced by direct sequencing of genomic DNA to detect genetic changes. The ability to perform such studies depends equally on the development of high-throughput and economical genotyping methods. Currently, basically for every disease whose origen is still unknown, genetic approaches are available which could be pedigree-dependent or -independent with the capacity to elucidate fundamental disease mechanisms. Computer algorithms and programs for linkage analysis have formed the foundation for many disease gene detection projects, similarly databases of clinical findings have been widely used to support diagnostic decisions in dysmorphology and general human disease. For every disease type, genome sequence variations, particularly single nucleotide polymorphisms are mapped by comparing the genetic makeup of case and control groups. Methods that predict the effects of polymorphisms on protein stability are useful for the identification of possible disease associations, whereas structural effects can be assessed using methods to predict stability changes in proteins using sequence and/or structural information.

Genetic heterogeneity in Pakistani microcephaly families.

Autosomal recessive primary microcephaly (MCPH) is caused by mutations in at least eight different genes involved either in cell division or DNA repair. Most mutations are identified in consanguine families from Pakistan, Iran and India. To further assess their genetic heterogeneity and mutational spectra, we have analyzed 57 consanguine Pakistani MCPH families. In 34 MCPH families, we detected linkage to five out of the eight well-characterized disease loci and identified mutations in 27 families, leaving seven families without mutations in the coding exons of the presumably underlying MCPH genes. In the MCPH cohort 23 families could not be linked to any of the known loci, pointing to remarkable locus heterogeneity. The majority of mutations were found in ASPM followed by WDR62, CENPJ, CEP152 and MCPH1. One ASPM mutation (p.Trp1326*) was found in as many as eight families suggesting a Pakistani founder mutation. One third of the families were linked to ASPM followed by WDR62 confirming previous data. We identified three novel ASPM mutations, four novel WDR62 mutations, one novel MCPH1 mutation and two novel CEP152 mutations. CEP152 mutations have not been described before in the Pakistani population.

Prevention of beta-thalassemia in a large Pakistani family through cascade testing.

BACKGROUND: We report cascade testing of a large Pakistani family for beta-thalassemia alleles. The family was still practicing consanguineous marriages and was at risk of having more affected births. OBJECTIVE: The objective of this study was to show that identification of disease carriers in families with index cases in order to create awareness about disease and provide genetic counseling would result in reduction of the frequency of beta-thalassemia in Pakistan. METHODS: In this large family with an index case, 27 available living members were tested for beta-thalassemia. Carriers of the disease were detected by measuring hemoglobin indices, and amplification refractory mutation system polymerase chain reaction was used for mutation analysis. Genetic counseling was provided to members of this family. RESULTS: There were already 3 marriages between the carrier members and 1 between a carrier and noncarrier in this large family; 12 (44.4%) members were found to carry the mutant gene, representing a very high carrier rate compared to the 5.4% carrier frequency of beta-thalassemia in the general population of Pakistan. The family was counseled for prevention of affected births. The initially reluctant family gradually became cooperative and seriously attended the genetic counseling sessions. CONCLUSION: Cascade testing is more practical than general population screening in a country with limited health facilities where consanguineous marriages are practiced. This report emphasizes the need of extensive testing within families with index cases to identify the carriers of beta-thalassemia in order to reduce disease occurrence through awareness and genetic counseling.

Clinical spectrum of Parkinson's disease from Pakistan.

INTRODUCTION: Parkinson's disease is an idiopathic disorder of the extrapyramidal system. It has a worldwide prevalence but data from developing countries is scanty. We describe the clinical spectrum of the disease from Pakistan, a developing country. METHODS: Patients with Parkinson's disease, over a period of 11 years, were identified by ICD-9 coding system of the hospital medical records. Demographical characteristics, clinical features, laboratory investigations and radiological investigations were recorded and analysed. RESULTS: A total of 80 patients were identified. 50 (63 percent) were males and 30 (37 percent) were females. Mean age of onset of the disease was 54 years. 47 (59 percent) patients had onset of illness during the sixth or seventh decade of life. Mean duration of illness at the time of presentation was five years. Rigidity, bradykinesia, tremors, hypomimia, primitive reflexes, difficulty in performing fine work and walking difficulty were the most common clinical features. 52 (65 percent) patients had stage I or II (Hoehn-Yahr staging) disease at the time of presentation. 56 (70 percent) patients had predominantly unilateral symptoms. 15 (19 percent) patients had cognitive impairment. Cognitive decline was more common in the elderly and in patients with disease duration of longer than ten years. CONCLUSION: Parkinson's disease is more common in males. Tremor, rigidity, walking difficulty, bradykinesia and difficulty in performing fine work are the commonest clinical features. Disease severity increases with duration of the disease. Cognitive impairment is not uncommon in these patients and is associated with disease duration and age of onset of the illness.

Stroke-associated pneumonia: microbiological data and outcome.

INTRODUCTION: Pneumonia is a common complication after acute stroke. It affects the outcome adversely. However, data regarding microbiology of stroke-associated pneumonia and its effect on outcome is scarce. METHODS: Stroke-associated pneumonia was identified through chart review of all ICD-9 identified adult stroke patients admitted to our hospital over a period of four years (1998-2001). The demographical, laboratory, radiological, microbiological data and outcome of patients with stroke-associated pneumonia were recorded and analysed. RESULTS: 443 patients with stroke were admitted over the four-year period and 102 (23 percent) had stroke-associated pneumonia. Their ages range from 28 to 100 (mean 64+/-14) years. 69 (68 percent) were men. Median length of stay was nine days compared to four days for all stroke patients. 68 (67 percent) patients manifested pneumonia within 48 hours and 34 (33 percent) after 48 hours of admission. Yield of tracheal aspirate cultures was 38 percent and that of chest radiographs was 25 percent. Pseudomonas aeruginosa and Staphylococcus aureus were the most common organisms (12 percent each) followed by Streptococcus pneumoniae and Klebsiella pneumoniae (4 percent each). Patients with infiltrates on chest radiographs were more likely to have positive tracheal aspirate cultures (p-value is 0.003). 35 patients (34 percent) expired during hospital stay. Positive chest radiographs and tracheal aspirates were independent predictors of prolonged hospital stay (p-value is less than 0.005). CONCLUSION: Pneumonia is a common medical complication of stroke. It is associated with a high mortality and prolongs the hospital stay. The yield of chest radiographs and tracheal aspirates is low. However, these are independent predictors of prolonged hospital stay. Pseudomonas aeruginosa and Staphylococcus aureus are most common organisms in stroke-associated pneumonia.

The neurochemical markers in cerebrospinal fluid to differentiate between aseptic and tuberculous meningitis.

In this study, the use of neurochemical markers in patients with aseptic and tuberculous meningitis has been investigated. The cerebrospinal fluid levels of amino acids, nitrite (a metabolite of nitric oxide), vitamin B12 and homocysteine were quantitated in both groups of patients. Among the amino acids, aspartic acid and glutamic acid both excitatory amino acid, GABA, glycine and tryptophan were all significantly increased in both patient groups whereas decreased level of taurine and increased level of phenylalanine were only found in patients with tuberculous meningitis. The levels of nitrite and its precursor arginine were significantly higher in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. A significantly increased homocysteine level and a decreased level of vitamin B12 were found only in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. This indicates that patients with tuberculous meningitis are particularly prone to vitamin B12 deficiency resulting into increased level of HC, and involvement of free radical showing the importance of these biological markers for promoting the possibility for the design of therapeutic approach.

Cerebral involvement in benign tertian malaria.

Although Plasmodium vivax usually causes benign uncomplicated malaria, it can occasionally result in severe disease with life-threatening, end-organ involvement generally seen with falciparum malaria. We report a case of cerebral malaria caused by P. vivax and review the literature on this subject.

Acetazolamide therapy improves action myoclonus in Ramsay Hunt Syndrome.

Acetazolamide treatment significantly improves action myoclonus in Ramsay Hunt Syndrome. A family with two brothers and a sister, and a sporadic case with Ramsay Hunt Syndrome and uncontrollable action myoclonus, are described where addition of oral acetazolamide lead to marked improvement in their action myoclonus.

Cytomegalovirus-associated transverse myelitis in a non-immunocompromised patient.

Cytomegalovirus-associated transverse myelitis (TM) is rare in immunocompetent patients. We report here the case of a 24-year-old woman who developed acute transverse myelitis with cerebrospinal fluid (CSF) findings, suggesting a central nervous system (CNS) infection, and a positive CMV serology.

Pattern of sex steroids secretion and their relationship with embryo yield in Jersey cows superovulated with PMSG.

The levels of progesterone and estrogen secretion were studied in relationship to the superovulatory response in Jersey cows. Progesterone and estrogen concentrations were measured in superovulated Jersey cows with the objective of correlating the patterns of steroid secretion with embryo yield and quality. Pregnant mare serum gonadotropin (PMSG) was used in combination with prostaglandin F(2) alpha analogue to induce superovulation in 18 multiparous, cyclic cows. Serum progesterone and estradiol levels from cows which exhibited estrus within 24 to 48 h after prostaglandin administration (n=13) were used to estimate the superovulatory response. Sex steroid concentrations at the day of estrus (Day 0) was a strong indicator of embryo yield. Progesterone was negatively (r=-0.56) and estrogen positively (r=0.80) correlated to the number of embryos collected. Dramatic increase in progesterone from Day 0 to Day 7 was a significant indicator of embryo yield. A higher rise of estrogen in the follicular phase was an indicator of a larger number of growing follicles and, consequently, better superovulatory response. Nonresponding animals did not show any significant change in the hormonal profile from the day of PMSG treatment to the day of embryo collection. The estimation of progesterone and estradiol concentrations, simultaneously, gave a more objective prediction of embryo yield.

Variation in cerebrospinal fluid levels of neuropeptide Y, cholecystokinin and substance P in patients with neurological disorders.

Neuropeptide Y, cholecystokinin (tetra- and octasulphated peptides) and substance P were measured in lumbar cerebrospinal fluid obtained from patients with various neurologic disorders such as Parkinson's disease, cerebrovascular disorders, multiple sclerosis, tuberculous meningitis and aseptic meningitis. These results are statistically compared with healthy results. The results accumulated showed that the data collected can provide the vital information necessary for designing drug therapy.

Ischemic stroke in young adults of South Asia.

OBJECTIVE: To study the risk factors, etiology and management of ischemic stroke in young adults in a South Asian population. METHODS: Retrospective study conducted at a large tertiary hospital in Karachi. One hundred and eighteen patients between fifteen and forty-five years admitted over a five year period with a diagnosis of ischemic infarct constituted the study population. The study variables included the full clinical spectrum, spanning historical, laboratory, radiological and outcome parameters. RESULTS: Forty-three percent of patients were hypertensive and 30% were diabetic. The combination of diabetes and hypertension was found in 19.5% of patients, intra-cranial and carotid artherosclerosis in 22% and embolism in 11% of the cases. The in-hospital mortality was 11%. The outcome was excellent in 27%, good in 50% and poor in 23% of patients. CONCLUSION: The risk factors for artherosclerosis and the contribution of intra and extra-cranial artherosclerosis were found to be much higher than those from the Western Hemisphere.

Moyamoya disease: an elusive diagnosis.

OBJECTIVE AND BACKGROUND: Moyamoya disease is an idiopathic vasculopathy of circle of Willis. Stroke is a common presentation. We describe clinical and radiological features of moyamoya disease in four patients. SETTING: Tertiary care center in a metropolitan city. MATERIALS AND METHODS: Patients with moyamoya disease were identified by ICD-9 coding system of the hospital medical records. Demographic characteristics, clinical features/presentation, laboratory investigations and radiological investigations were recorded and analyzed. RESULTS: Four patients (three children and one adult) presented with hemiparesis. One patient expired during hospital stay. Two patients underwent surgery (synangiosis). EEGs of three patients showed focal abnormalities. Neuroimaging showed ischemic infarctions in all patients. Findings on four vessel digital substraction angiography and/or magnetic resonance angiography were compatible with moyamoya disease. CONCLUSION: Moyamoya disease should be considered in all young patients, especially children, presenting with stroke.

Carotid Doppler ultrasonography in young stroke patients.

BACKGROUND: The present study focuses on the role of carotid doppler ultrasonography (CDUS) in the diagnosis and management of carotid stenosis in young stroke patients. METHODS: The findings of carotid doppler in 45 ischemic stroke patients between 15-45 years of age were reviewed retrospectively. The variables of interest for this study included risk factors for atherosclerotic disease, primary abnormality detected on carotid doppler ultrasonography (ulceration vs. stenosis), degree of stenosis and the type of plaque (soft vs. calcified). RESULTS: The prevalence of hypertension and diabetes was 50% and 35% respectively. The rate of carotid stenosis in the study population was found to be 31%. The degree of stenosis was mild in 35% and moderate in 21%. High-grade stenosis was found in 21% of patients. The plaque was soft in the majority of cases (43%). CONCLUSION: The proportion of carotid stenosis in young stroke patients was relatively high compared with previous studies. This may be due to an increase in the risk factors for atherosclerotic disease in developing countries.

Effect of chemotherapy on circulating steroid hormone levels in postoperative premenopausal breast cancer patients.

Serum levels of 17-beta oestradiol, testosterone and progesterone were determined in postoperative premenopausal breast cancer patients. In patients receiving chemotherapy circulating 17-beta oestradiol values decreased significantly compared to control group during the sampling/drug regimens employed. Among the control group, however, the oestradiol levels remained high throughout the sampling period. Testosterone levels in patients were also significantly low compared to control group throughout the sampling regimen up to 28 days. In contrast the levels of progesterone in patients were elevated and remained high compared to the corresponding controls. A positive correlation was found between the drop in serum oestradiol and testosterone levels following the initiation of chemotherapy and the regression of the tumour size. Steroid hormone levels in the serum of breast cancer patients receiving chemotherapy can serve as clinical tools to monitor the progress of the disease and response to therapy.

Should carotid endarterectomy be performed for symptomatic carotid stenosis in Pakistan?

OBJECTIVE: The risk of stroke and death associated with carotid endarterctomy is operator dependant. Data regarding risks of this procedure are not available in Pakistan and therefore it is difficult to make accurate risk benefit analysis for individual patients. Our objective was to determine safety of carotid endarterectomy at an academic tertiary care center in Pakistan. METHODS: Patients who underwent carotid endarterectomy (CEA) at our hospital during a ten-year period were identified through ICD-9 coding system of the hospital medical records. Demographic features, associated medical problems and immediate postoperative complications were recorded and analyzed. RESULTS: Sixty-three carotid endarterectomies were performed on 59 patients. Ages range from 43 to 80 (mean 61 +/- 8) years; 53 were male and 10 were female. Common associated diseases among these patients were hypertension; 38 (64.4%), ischemic heart disease; 26 (44%), diabetes mellitus; 24 (40.7%), dyslipidemia; 19 (32.2%) and renal insufficiency; 13 (22%). Most common complication was neuropraxia (transient neuropathy); 5 (7.9%), followed by pneumonia and stroke; each in 3 (4.8%) patients. None of the strokes related to the surgical procedure were disabling. Two of the patients who had stroke, recovered fully within 17 weeks and one recovered partly but was independent in all daily activities of living (ADLs). One patient died following simultaneous coronary artery bypass graft (CABG) and CEA. The risk of stroke or death for patients undergoing CEA was high with simultaneous CABG (3/11, 27%) and low for patients undergoing CEA alone (1/52, 2%). CONCLUSION: Carotid endarterectomy is a safe procedure in patients with symptomatic carotid stenosis at our hospital and should be performed, when indicated.

Impact of the long term supply of iodized salt to the endemic goitre area.

The study was designed to assess the impact of the supplementation levels of iodine in salt supplied since the last 12 years to Gilgit and Hunza, an endemic goitre area of Pakistan. The overall prevalence of visible goitre is reduced from 61.36% to 4.68%. Results of urinary excretion of iodine (UEI) indicate severe to mild iodine deficiency among 70.41% of the randomly surveyed households. Severely deficient are 3%, moderate 29.54% and mild 37.87%, criteria of UEI being less than 2.0 micrograms/dl, 2-5 micrograms/dl and 5-10 micrograms/dl respectively. Levels of iodine supplementation in 267 iodized salt samples at production (n = 128) and consumption (n = 139) points are compared with a mean +/- SD are 70.86 +/- 29.73 ppm and 37.24 +/- 20.47 ppm respectively, representing 566.8 +/- 237.8 micrograms and 297.9 +/- 163.7 micrograms of iodine per 8.0 gram of salt. It is suggested to replace common salt with iodized salt in the goitre area to ensure the use by all households and quality control measures for iodination of salt should strictly be adhered so that uniform and consistent supply of iodine be ensured. The magnitude of contributory factors other than iodine deficiency, i.e., environmental and hereditary should be monitored and considered when levels of iodine supplementation are adjusted.