Synchronization of ovulation and fertility in weaned sows treated with intravaginal triptorelin is influenced by timing of administration and follicle size.

A 100 µg dose of triptorelin was tested for synchronizing ovulation in sows. In Experiment 1, conducted in April through June, sows (n = 125) were assigned to Control (untreated), TG-96 (Triptorelin Gel (TG) given intravaginally at 96 h post-weaning), or TG-E (given intravaginally at estrus). To optimize AI timing, sows were inseminated at 2 and 26 h after estrus for Control and TG-E and at 8 and 32 h following TG-96. Ovulation by 48 h post-treatment tended to be affected by treatment (P = 0.08) and more (P < 0.05) TG-96 sows ovulated (57.9%) compared to Controls (34.2%), but TG-E (45.1%) did not differ (P > 0.10). Duration of estrus was reduced (P < 0.005) in TG-96 (51 h) and TG-E (58 h) compared to Controls (65 h). There was no treatment effect on farrowing rate (71%) or total born (10.4). Average follicle size <6.5 mm at 96 h after weaning was associated with reduced (P < 0.01) estrus, ovulation and farrowing rate. Experiment 2 was conducted in August through September using 503 weaned sows. The TG-96 treatment reduced duration of estrus (P = 0.03), but treatment did not affect estrus expression, farrowing rate or total pigs born. In conclusion, use of a 100 µg dose of triptorelin intravaginally at 96 h or at estrus advanced ovulation and when used with timed insemination, resulted in similar farrowing rates and litter sizes comparable to sows mated based on estrus. However, ovulation induction and timed AI success may benefit from an approach that ensures sows have adequate follicle development at time of treatment.

Endocrine, ovulatory and reproductive characteristics of sows treated with an intravaginal GnRH agonist.

The present study examined the effectiveness of intravaginal administration of a GnRH agonist, triptorelin, on the induction of the preovulatory LH surge; synchronization of time of ovulation; and reproductive characteristics in weaned sows. Sows were given 100microg of triptorelin in 0.9, 1.2, or 1.5% methylcellulose gel intravaginally 96h after weaning and then bred at 8 and 32h post-treatment. Untreated sows inseminated once each day of estrus served as the positive controls, while females treated with only the methyl cellulose gel and also bred once each day of estrus were negative controls. Sows treated intravaginally with triptorelin exhibited preovulatory LH surges with magnitudes comparable to those that occurred spontaneously in the negative controls. Preovulatory LH surges were initiated over a narrow and well-defined time interval that occurred 4-12h after treatment in sows receiving triptorelin in 1.2 or 1.5% methyl cellulose gel. As a result, the majority of the sows in these two treatments had ovulations within a 12h time frame 36-48h after treatment. In contrast, both the LH surge and ovulation occurred over extended periods of time after weaning in negative controls and sows given triptorelin in 0.9% methylcellulose gel. Farrowing rates and litter size were similar between untreated controls and triptorelin-treated sows that were bred with two fixed timed inseminations. Insemination of sows induced to have ovulations and bred at least once while not in estrus did not have any overt negative effects on reproductive characteristics. These results demonstrate that 100microg of triptorelin administered intravaginally in a least 1.2% methyl cellulose gel induced a normal preovulatory LH surge and synchronized time of ovulation in weaned sows. Furthermore, there were no obvious changes in reproductive performance when these sows were bred with two fixed time inseminations regardless of whether they exhibited a standing reflex.

[Salivary stones and stenosis. A comprehensive classification]. / Lithiases et sténoses salivaires. Une classification pratique des pathologies non tumorales.

INTRODUCTION: Sialendoscopy and sialo-MRI enable diagnosis of salivary gland obstructive pathologies, such as lithiasis, stenosis and dilatations. Therefore, a classification of these pathologies is needed, allowing large series comparisons, for better diagnosis and treatment of salivary pathologies. MATERIAL AND METHODS: With help from people from the European Sialendoscopy Training Center (ESTC), the results of sialographies, sialoMRI and sialendoscopies, a comprehensive classification of obstructive salivary pathologies is described, based on the absence or presence of lithiasis (L), stenosis (S) and dilatation (D) ("LSD" classification). DISCUSSION: It appears that a classification of salivary gland obstructive pathologies should be described. We hope it will be widely used and of course criticized to be improved and to compare the results of salivary gland diagnostic methods, such as sialography and sialendoscopy and also the results and indications for salivary gland therapeutic methods, such as lithotripsy, sialendoscopy and/or open surgery.

Salivary stones and stenosis. A comprehensive classification.

INTRODUCTION: Sialendoscopy and sialoMRI enables diagnosis of salivary gland obstructive pathologies, such as lithiasis, stenosis, and dilatations. Therefore, a classification of these pathologies is needed, allowing large series comparisons, for better diagnosis and treatment of salivary pathologies. MATERIAL AND METHODS: With help from people from the European Sialendoscopy Training Center (ESTC), the results of sialographies, sialoMRI and sialendoscopies, a comprehensive classification of obstructive salivary pathologies is described, based on the absence or presence of lithiasis (L), stenosis (S), and dilatation (D) ("LSD" classification). DISCUSSION: It appears that a classification of salivary gland obstructive pathologies should be described. We hope it will be widely used and of course criticized to be improved and to compare the results of salivary gland diagnostic methods, such as sialography and sialendoscopy, and also the results and indications for salivary gland therapeutic methods, such as lithotripsy, sialendoscopy, and/or open surgery.

Effects of nucleotide supplementation in milk replacer on small intestinal absorptive capacity in dairy calves.

Milk replacer was supplemented with nucleotides and fed to dairy calves from birth through weaning to examine the potential for enhancing recovery of small intestinal function after enteric infection. Three treatments of 23 calves each were fed milk replacer (10% body weight/d) supplemented with no nucleotides (C), purified nucleotides (N), or nucleotides from an extract of Saccharomyces cerevisiae (S). Average daily gain, health scores, fecal dry matter, and fecal bacteria were monitored, and blood was analyzed for packed cell volume, glucose, blood urea nitrogen (BUN), and creatinine. Calves were monitored twice daily for fecal score, and 48 h after increased fecal fluidity was recorded, intestinal function was evaluated by measuring absorption of orally administered xylose (0.5 g/kg of body weight). Packed cell volume of blood was greater for treatment N for wk 2 and 5 compared with other treatment groups. Four calves per treatment were killed, and intestinal tissue was evaluated for morphology, enzyme activities, and nucleoside transporter mRNA expression. Treatment S calves had increased abundance of nucleoside transporter mRNA, numerically longer villi, and lower alkaline phosphatase than other groups. Growth measurements and plasma concentrations of glucose, BUN, creatinine, and IgG were not different between treatments; however, BUN-to-creatinine ratio was higher for treatment N, possibly indicating decreased kidney function. There were also no treatment effects on fecal dry matter and fecal bacteria population. However, N-treated calves had the highest detrimental and lowest beneficial bacteria overall, indicating an unfavorable intestinal environment. Supplementation of purified nucleotides did not improve intestinal morphology or function and resulted in higher fecal water loss and calf dehydration. Supplementation of nucleotides derived from yeast tended to increase calf intestinal function, provide a more beneficial intestinal environment, and improve intestinal morphology.

Effects of dexamethasone on mRNA abundance of nuclear receptors and hepatic nuclear receptor target genes in neonatal calves.

Hepatic nuclear receptors (NR), particularly constitutive androstane receptor (CAR) and pregnane X receptor (PXR), are involved in the coordinated transcriptional control of genes that encode proteins involved in the metabolism and detoxification of xeno- and endobiotics. A broad spectrum of metabolic processes are mediated by NR acting in concert with ligands such as glucocorticoids. This study examined the role of dexamethasone on hepatic mRNA expression of CAR, PXR and several NR target genes. Twenty-eight male calves were allotted to one of four treatment groups in a 2 x 2 arrangement of treatments: feed source (colostrum or milk-based formula) and glucocorticoid administration (twice daily intramuscular dexamethasone). Liver biopsies were obtained at 5 days of age. Real-time reverse transcription polymerase chain reaction was used to quantify mRNA abundances. No effects of feed source on mRNA abundances were observed. For the NR examined, mRNA abundance of both CAR and PXR in dexamethasone-treated calves was lower (p < 0.05) by 39% and 40%, respectively, than in control calves. Abundance of NR target genes exhibited a mixed response. SULT1A1 mRNA abundance was 39% higher (p < 0.05) in dexamethasone-treated calves compared with control calves. mRNA abundance of CYP2C8 tended also to be higher (+44%; p = 0.053) after dexamethasone treatment. No significant treatment effects (p > 0.10) were observed for mRNA abundances of CYP3A4, CYP2E1, SULT2A1, UGT1A1 or cytochrome P450 reductase (CPR). In conclusion, an enhanced glucocorticoid status, induced by pharmacological amounts of dexamethasone, had differential and in part unexpected effects on NR and NR target systems in 5-day-old calves. Part of the unexpected responses may be due the immaturity of NR and NR receptor target systems.

Insulin-like growth factor type-1 receptor down-regulation associated with dwarfism in Holstein calves.

Perturbations in endocrine functions can impact normal growth. Endocrine traits were studied in three dwarf calves exhibiting retarded but proportionate growth and four phenotypically normal half-siblings, sired by the same bull, and four unrelated control calves. Plasma 3,5,3'-triiodothyronine and thyroxine concentrations in dwarfs and half-siblings were in the physiological range and responded normally to injected thyroid-releasing hormone. Plasma glucagon concentrations were different (dwarfs, controls>half-siblings; P<0.05). Plasma growth hormone (GH), insulin-like growth factor-1 (IGF-1) and insulin concentrations in the three groups during an 8-h period were similar, but integrated GH concentrations (areas under concentration curves) were different (dwarfs>controls, P<0.02; half-siblings>controls, P=0.08). Responses of GH to xylazine and to a GH-releasing-factor analogue were similar in dwarfs and half-siblings. Relative gene expression of IGF-1, IGF-2, GH receptor (GHR), insulin receptor, IGF-1 type-1 and -2 receptors (IGF-1R, IGF-2R), and IGF binding proteins were measured in liver and anconeus muscle. GHR mRNA levels were different in liver (dwarfs

Nuclear receptor and nuclear receptor target gene messenger ribonucleic acid levels at different sites of the gastrointestinal tract and in liver of healthy dogs.

Nuclear receptors (NR) are ligand-activated transcription factors that regulate different metabolic pathways by influencing the expression of target genes. The current study examined mRNA abundance of NR and NR target genes at different sites of the gastrointestinal tract (GIT) and the liver of healthy dogs (Beagles; n = 11). Samples of GIT and liver were collected postmortem and homogenized, total RNA was extracted and reverse transcribed, and gene expression was quantified by real-time reverse-transcription PCR relative to the mean of 3 housekeeping genes (beta-actin, glyceraldehyde-3-phosphate dehydrogenase, and ubi-quitin). Differences were observed (P < or = 0.05) in the mRNA abundance among stomach (St), duodenum (Du), jejunum (Je), ileum (Il), and colon (Col) for NR [pregnane X receptor (Du, Je > Il, Col > St), peroxisome proliferator-associated receptor gamma (St, Du, Col > Je, Il), constitutive androstane receptor (Je, Du > Il, Col), and retinoid x receptor alpha (Du > Il)] and NR target genes [glutathione-S-transferase A3-3 (Du > Je > St, Il; St > Col), phenol-sulfating phenol sulfotransferase 1A1 (Du, Je > Il, St; Col > St), cytochrome P450 3A12 (Du, Je > St, Il, Col), multiple drug resistance gene 1 (Du, Je, Il, Col > St), multiple drug resistance-associated protein 2 (Je, Du > Il > St, Col), multiple drug resistance-associated protein 3 (Col > St > Il; Du > Je, Il; St > Il), NR corepressor 2 (St > Il, Col), and cytochrome P450 reductase (St, Du, Je > Il, Col)], but not for peroxisome proliferator-associated receptor alpha. Differences (P > 0.05) in mRNA abundance in the liver relative to the GIT were also observed. In conclusion, the presence of numerous differences in expression of NR and NR target genes in different parts of the GIT and in liver of healthy dogs may be associated with location-specific functions and regulation of GIT regions.

Nuclear receptor and target gene mRNA abundance in duodenum and colon of dogs with chronic enteropathies.

Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR) and peroxisome proliferator-associated receptors alpha and gamma (PPARalpha, PPARgamma) are mediators of inflammation and may be involved in inflammatory bowel disease (IBD) and food responsive diarrhea (FRD) of dogs. The present study compared mRNA abundance of NR and NR target genes [multi drug-resistance gene-1 (MDR1), multiple drug-resistance-associated proteins (MRD2, MRD3), cytochrome P450 (CYP3A12), phenol-sulfating phenol sulfotransferase (SULT1A1) and glutathione-S-transferase (GST A3-3)] in biopsies obtained from duodenum and colon of dogs with IBD and FRD and healthy control dogs (CON; n=7 per group). Upon first presentation of dogs, mRNA levels of PPARalpha, PPARgamma, CAR, PXR and RXRalpha in duodenum as well as PPARgamma, CAR, PXR and RXRalpha in colon were not different among groups (P>0.10). Although mRNA abundance of PPARalpha in colon of dogs with FRD was similar in both IBD and CON (P>0.10), PPARalpha mRNA abundance was higher in IBD than CON (P<0.05). Levels of mRNA of MDR1 in duodenum were higher in FRD than IBD (P<0.05) or CON (P<0.001). Compared with CON, abundances of mRNA for MRP2, CYP3A12 and SULT1A1 were higher in both FRD and IBD than CON (P<0.05). Differences in mRNA levels of PPARalpha and MRP2 in colon and MDR1, MRP2, CYP3A12 and SULT1A1 in duodenum may be indicative for enteropathy in FRD and (or) IBD dogs relative to healthy dogs. More importantly, increased expression of MDR1 in FRD relative to IBD in duodenum may be a useful diagnostic marker to distinguish dogs with FRD from dogs with IBD.

Ontogeny of mRNA abundance of nuclear receptors and nuclear receptor target genes in young cattle.

After birth the development of appropriate detoxification mechanisms is important. Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR), peroxisome proliferator-activated receptor-alpha (PPARalpha), retinoid receptors (RAR, RXR), and NR target genes are involved in the detoxification of exogenous and endogenous substances. We quantified abundances of hepatic mRNA of NR and several NR target genes (cytochromes, CYP; cytochrome P450 reductase, CPR; UDP-glucuronosyl transferase, UDP) in calves at different ages. Gene expression was quantified by real-time RT-PCR. Abundance of mRNA of CAR and PXR increased from low levels at birth in pre-term calves (P0) and full-term calves (F0) to higher levels in 5-day-old calves (F5) and in 159-day-old veal calves (F159), whereas mRNA levels of PPARalpha did not exhibit significant ontogenetic changes. RARbeta mRNA levels were higher in F5 and F159 than in F0, whereas no age differences were observed for RARalpha levels. Levels of RXRalpha and RXRbeta mRNA were lower in F5 than in P0 and F0. Abundance of CYP2C8 and CYP3A4 increased from low levels in P0 and F0 to higher levels in F5 and to highest levels in F159. Abundance of CPR was transiently decreased in F0 and F5 calves. Levels of UGT1A1 mRNA increased from low levels in P0 and F0 to maximal level in F5 and F159. In conclusion, mRNA levels of NR and NR target genes exhibited ontogenetic changes that are likely of importance for handling of xeno- and endobiotics with increasing age.

Expression of nuclear receptor and target genes in liver and intestine of neonatal calves fed colostrum and vitamin A.

Nuclear receptors (NR), including retinoic acid and retinoid X receptors (RAR, RXR), pregnane X receptor (PXR), constitutive androstane receptor, and peroxisome proliferator-activated receptor (PPARalpha) modify the expression of other genes, such as cytochrome p450 enzymes (CYP), sulfotransferases (SULT), and UDP glucuronosyl transferases (UGT). Nuclear receptor expression is influenced by exposure to ligands (e.g., vitamin A). We tested the hypothesis that vitamin A feeding influences the expression of hepatic and intestinal NR and their target genes and that colostrum or formula feeding influence these traits differently. Calves (n = 7/ group) were fed colostrum (CO) or a milk-based formula with or without vitamin A (FA, FO, respectively) for 4 d and were euthanized on d 5, followed immediately by tissue collection. Thereafter, RNA was extracted and gene expression quantified by real-time reverse transcription-polymerase chain reaction. Expression relative to housekeeping genes of mRNA was profiled for NR, CYP, SULT, and UGT enzymes. Hepatic mRNA levels of RARbeta and CYP26 were higher in FA than FO cows; expression of CYP2E1, CYP2C8, CYP26, and UGT1A1 was higher in CO than FO cows; and expression of CYP2E1, UGT1A1, and p450 reductase was higher in CO than FA. In colon tissue, abundance of RXRalpha mRNA was lower in FO than CO, and CYP2B6 expression was lower in FO than in CO and FA. In jejunal tissue, there were no significant differences in gene expression among groups. In conclusion, effects of vitamin A feeding were limited, but colostrum feeding had several selective effects on expression of nuclear receptors and target genes.

Gene polymorphisms in detoxification enzymes as susceptibility factor for head and neck cancer?

OBJECTIVE: Impaired detoxification of carcinogens found in tobacco smoke appears to increase the risk for tobacco associated cancer. The objective of this study was to investigate concomitant polymorphisms in genes encoding for various detoxification enzymes in patients with head and neck squamous cell carcinoma (HNSCC). METHODS: In 187 patients with HNSCC and in 139 healthy control subjects, the polymorphisms of cytochrome P450 1A1 (CYP1A1), cytochrome P450 2D6 (CYP2D6), and glutathione S-transferase mu1 and Theta (GSTM1, GSTT1) were detected by polymerase chain reaction. RESULTS: No significant association were identified between CYP1A1 and CYP2D6 gene polymorphisms and HNSCC. Patients with laryngeal cancer revealed the GSTM1 null genotype more frequently than did the control subjects (P < 0.05). The coincidence of GSTM1 and GSTT1 null genotype was found twice as great in patients as in control subjects (P < 0.05). CONCLUSIONS: It is assumed that detoxification enzymes are functionally redundant and only the simultaneous deficiency of several detoxification enzymes increase the risk for HNSCC in alcohol- and tobacco-exposed individuals.

GSTM1 enzyme concentration and enzyme activity in correlation to the genotype of detoxification enzymes in squamous cell carcinoma of the oral cavity.

BACKGROUND: Differences in genotype and phenotype of detoxification genes could be one reason for conflicting results in studies dealing with gene polymorphisms as susceptibility factors for tobacco associated cancer. OBJECTIVES: The objective of this study was to investigate gene polymorphisms of detoxification enzymes and to determine whether the enzyme concentration and activity of glutathione S transferase microliter 1 correlates with the genotype in patients with cancer of the oral cavity. MATERIAL AND METHODS: In 73 cancer patients and 136 matched healthy controls, the polymorphisms of glutathione S-transferase mu1 and theta (GSTM/GSTT), cytochrome p450 1A1 and CYP2D6 were detected. Simultaneously, GSTM1 protein concentration and total GSTM1-activity were determined. RESULTS: Only the coincidence of GSTM1 and GSTT null genotype was associated with oral cavity cancer. GSTM1 protein concentration and enzyme activity in null-genotype patients was significantly lower than in GSTM1-allele-carrier. But the enzyme concentration did not correlate with the activity. CONCLUSION: We assume that detoxification enzymes are functionally redundant and that only the simultaneous deficiency of several detoxification enzymes increases the risk for oral cancer.

11Beta-hydroxysteroid dehydrogenase 1 expression in squamous cell carcinomas of the head and neck.

11Beta-hydroxysteroid dehydrogenase 1 (11beta-HSD1) has been identified as a major detoxification enzyme of one of the most potent tobacco smoke-derived carcinogens, NNK. If not metabolized by 11beta-HSD1, activation of NNK by cytochrome p450 mono-oxidase 2D6 (CYP2D6) results in an electrophile intermediate responsible for DNA damage. Interindividual variability in the expression of 11beta-HSD1 and CYP2D6 has been found to influence the susceptibility to lung cancer. The aim of this study was to compare 11beta-HSD1 mRNA expression and CYP2D6 metabolizer status in pharyngeal tissues of patients with oropharyngeal carcinoma and controls. In 20 patients with oropharyngeal cancer and 15 non-smoking controls, the 11beta-HSD1 mRNA expression was assessed with RT-PCR. The frequency of genetic polymorphisms of the CYP2D6 gene was assessed using RFLP. It was found that 11beta-HSD1 mRNA is expressed in human pharyngeal mucosa. It is upregulated in mucosa exposed to tobacco smoke. In tumour tissues, 11beta-HSD1 expression was significantly lower than in non-affected mucosa. The frequency distribution of CYP2D6 gene polymorphisms was similar in patients and controls. Chronic tobacco abuse results in 11beta-HSD1 enzyme induction. A reduction of 11beta-HSD1 expression in tumour tissues could be a consequence of malignantly transformed cells. It remains unclear if the lower 11beta-HSD1 expression gives rise to an increased rate of additional mutations.

A comparative approach to structure-function studies of mammalian aromatases.

To date, structure--function studies of aromatase cytochrome P450 (P450arom) have been advanced by point mutation analyses utilizing almost exclusively the human enzyme, in conjunction with computer-generated models of the three-dimensional form of the enzyme based on prokaryotic cytochromes P450. Recent studies have identified duplicated isozymes of porcine P450arom, the gonadal and placental forms of which appear to differ substantially in substrate utilization and inhibitor sensitivity. We present a comparative approach to define regions of P450arom responsible for specific functional characteristics using complimentary DNAs encoding the porcine isozymes. Constructs encoding the native and chimeric porcine and human P450arom enzymes were transiently expressed and activity was assessed using the tritiated water assay. Sensitivity to inhibition by the imidazole etomidate was investigated, and P450arom expression was assessed by immunoblot analysis. All constructs yielded active P450arom, suggesting that exchanging entire structural elements does not preclude catalytic function. The activity of the gonadal isozyme was shown to be inhibited by etomidate at concentrations 185 and 300-fold lower than those required to induce a similar inhibition of the placental and human enzymes, respectively. In contrast, there was only a two-fold difference in the sensitivity of the gonadal and placental isozymes to inhibition by CGS16949A. Analysis of chimeric constructs indicated that the sensitivity to etomidate was associated with residues in the B, B' and C helices of the gonadal P450arom encompassing only one of six putative substrate recognition sites. Additionally, sensitivity to etomidate was not correlated with enzyme activity among the chimeric enzymes. Therefore, it appears that residues of the porcine gonadal P450arom that are responsible for etomidate binding may be distinct from those involved in substrate recognition and metabolism. These data support the notion that a comparative approach employing the use of chimeric enzymes provides a useful tool in directing point mutational analysis to determine residues important in inhibitor and perhaps substrate recognition of P450 enzymes such as P450arom. These studies are currently in progress.

[GSTM1 gene polymorphism in patients with head and neck tumors]. / GSTM1-Genpolymorphismus bei Patienten mit Kopf-Hals-Tumoren.

BACKGROUND: Glutathione S-Transferase mu (GSTM) is a phase II detoxification enzyme, which rapidly detoxifies carcinogens found in tobacco smoke. The prevalence of this polymorphism is about 50% in the caucasian population. The lack of GSTM1 has been linked with an increased susceptibility of smoking related cancers. A homozygote deletion of the GSTM-gene results in a missing gene product. The objective of this study was to investigate the frequency of the GSTM1 null genotype in squamous cell carcinoma of head and neck, especially the larynx and hypopharynx and to analyse the occurrence with respect to certain anatomical sites of cancer. MATERIAL AND METHODS: The GSTM1 genotypes of 83 patients with head and neck cancers and 60 healthy controls were determined by polymerase chain reaction (PCR) using blood leukocyte DNA. The presence or absence of the PCR-product after electrophoretic separation in an 2.0% agarose gel revealed the positive or negative genotype. RESULTS: The absence of the GSTM1 gene (null genotype) was found in 64% of all head and neck cancer patients and in 48% of the healthy controls (p < 0.05). Separating for cancer site, the null genotype was found in 44% of patients with hypopharyngeal cancer and in 78% of patients with laryngeal cancer (p < 0.05). The protein concentration of GSTM-enzyme correlated with the genotype. CONCLUSIONS: The results suggest that GSTM1 deficiency predisposes to head and neck cancer, especially to cancer of the larynx, which is particularly exposed to tobacco smoke carcinogens.

Characterization of an alphoid subfamily located near p-arm sequences on human chromosome 22.

The centromeric heterochromatin of all human chromosomes is composed of tandemly repeated alpha satellite DNA. Here we describe another alphoid subfamily that maps to human chromosome 22 as determined by FISH. The alphoid sequences were isolated from three YAC-clones carrying DNA from the pericentromeric region of the short arm of human chromosome 22 and limited amounts of alphoid DNA. This property enabled us to map the members of the subfamily to the border of the centromeric region and the short arm of the chromosome. The new alphoid subfamily may contribute to the closure of the gap remaining between the centromeric and short-arm maps of human chromosome 22.