Infant isolation and cohorting for preventing or reducing transmission of healthcare-associated infections in neonatal units.

BACKGROUND: Neonatal healthcare-associated infections (HAIs) result in increased morbidity and mortality, as well as increased healthcare costs. Patient isolation measures, i.e. single-room isolation or the cohorting of patients with similar infections, remain a recommended and commonly used practice for preventing horizontal spread of infections in the neonatal intensive care unit (NICU).  OBJECTIVES: Our primary objective was to assess the effect of single-room isolation or cohorting, or both for preventing transmission of HAIs or colonization with HAI-causing pathogens in newborn infants less than six months of age admitted to the neonatal intensive care unit (NICU). Our secondary objective was to assess the effect of single-room isolation or cohorting, or both on neonatal mortality and perceived or documented adverse effects in newborn infants admitted to the NICU.  SEARCH METHODS: We searched the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, Embase, CINAHL, the WHO ICTRP and ClinicalTrials.gov trials registries. There were no restrictions to date, language or publication type. We also checked the reference lists of studies identified for full-text review.  SELECTION CRITERIA: Types of studies: cluster-randomized or quasi-randomized trials at the level of the cluster (where clusters may be defined by NICU, hospital, ward, or other subunits of the hospital). We also included cross-over trials with a washout period of more than four months (arbitrarily defined). TYPES OF PARTICIPANTS: newborn infants less than six months of age in neonatal units that implemented patient isolation or cohorting as infection control measures to prevent HAIs. Types of interventions: patient isolation measures (single-room isolation or cohorting, or both of infants with similar colonization or infections) compared to routine isolation measures. TYPES OF OUTCOME MEASURES: the primary outcome was the rate of transmission of HAIs as estimated by the infection and colonization rates in the NICU. Secondary outcomes included all-cause mortality during hospital stay at 28 days of age, length of hospital stay, as well as potential adverse effects of isolation or cohorting measures, or both. DATA COLLECTION AND ANALYSIS: The standard methods of Cochrane Neonatal were used to identify studies and assess the methodological quality of eligible cluster-randomized trials. The certainty of the evidence was to be assessed by the GRADE method as evidence of high, moderate, low, or very low certainty. Infection and colonization rates were to be expressed as rate ratios for each trial and if appropriate for meta-analysis, the generic inverse variance method in RevMan was to be used. MAIN RESULTS: We did not identify any published or ongoing trials to include in the review. AUTHORS' CONCLUSIONS: The review found no evidence from randomized trials to either support or refute the use of patient isolation measures (single-room isolation or cohorting) in neonates with HAIs. Risks secondary to infection control measures need to be balanced against the benefits of decreasing horizontal transmission in the neonatal unit for optimal neonatal outcomes. There is an urgent need to research the effectiveness of patient isolation measures for preventing the transmission of HAIs in neonatal units. Well-designed trials randomizing clusters of units or hospitals to a type of patient isolation method intervention are warranted.

Novel Score to Predict Immunoglobulin Resistance in Kawasaki Disease.

To evaluate existing scoring systems and develop a new model to predict intravenous immunoglobulin (IVIG) resistance in patients with Kawasaki disease (KD). A retrospective cohort study performed between 2004 and 2017 identified 115 patients treated with IVIG for classic or incomplete KD. In our practice, IVIG resistance was defined as fever for > 24 h and patients were divided into responders and non-responders. A univariate analysis was performed to identify independent predictors of IVIG resistance. The predictors were combined into a new scoring system and compared with existing scoring systems. Sixty-five patients had classic KD and 50 had incomplete KD. Among the 115 patients, 80 (69.6%) responded and the remaining 35 were resistant (30.4%) to IVIG. Of the 35 resistant patients, 16 patients had incomplete KD. Hispanic children comprised 43% of our sample population. Coronary artery abnormalities developed in 14 of the 35 IVIG-resistant patients (39%). Univariate analysis showed that IVIG-resistant patients were older and present with lower platelets, potassium, and creatinine (P < 0.05). Multivariate logistic regression analysis used platelets, potassium, body surface area (BSA), and creatinine to devise the Las Vegas Scoring System (LVSS), which demonstrated a sensitivity of 76.2% and a specificity of 68.6%. Compared to published data, we observed a higher rate of IVIG resistance and coronary artery abnormalities in our patient population. The LVSS (using platelets, potassium, BSA, and creatinine) showed higher specificity and comparable sensitivity to other scoring systems devised to predict IVIG resistance.

Interventional Pain Treatments in the Management of Oncologic Patients With Thoracic Spinal Tumor-Related Pain: A Case Series.

BACKGROUND: Advanced tumors of the thoracic spine are difficult to treat and can lead to complex pain syndromes. Following conventional oncologic treatments, pharmacologic therapy may be insufficient to manage pain. Minimally invasive interventional procedures offer alternatives to treat malignant thoracic spinal pain. METHODS: Thirteen patients with metastatic disease and poorly controlled thoracic axial and/or radicular pain were identified via a retrospective chart review. Patients were either treated with radiation, surgery, chemotherapy, or a combination of these. Then, the patients were organized into groups based on their diagnoses, anatomical disease locations, symptoms, prior treatments, and interventional pain procedures offered. RESULTS: All cases of intercostal nerve, costotransverse junction, erector spinae plane, and paravertebral blocks resulted in pain relief without any reported complications. A patient who received a thoracic epidural injection had a complete resolution of pain when combined with radiation therapy 2 weeks after the injection. One patient who underwent repeat thoracic epidural injections eventually had an intrathecal pump placement, resulting in reduced opioid usage. Finally, 1 patient who received a thoracic medial branch block with a relief of thoracic axial pain reported greater pain relief with a medial branch nerve cryoablation. CONCLUSION: We propose a treatment algorithm to manage patients with thoracic spinal tumor-related pain. Interventional thoracic axial procedures may be safe and efficacious pain treatments for patients with cancer.

Genetic association of the glycine cleavage system genes and myelomeningocele.

BACKGROUND: Neural tube defects (NTDs) are one of the most common congenital birth defects, with myelomeningocele (MM) being the most severe form compatible with life. Recent studies show a link between mitochondrial folate one carbon metabolism and NTDs by means of the glycine cleavage system (GCS). We hypothesize that single nucleotide polymorphisms and novel variants in the coding regions of the GCS genes increase the risk for MM. METHODS: DNA was obtained from 96 subjects with MM born before the United States mandated folic acid fortification of grains in 1998. Primers were designed for polymerase chain reaction amplification and sequencing of all exons in the AMT gene, one of four genes in the GCS, followed by identification of single nucleotide polymorphisms and novel variants. An additional 252 MM subjects underwent whole exome sequencing to examine all four GCS genes (aminomethyltransferase, glycine dehydrogenase, glycine cleavage system protein-H, and dihydrolipoamide dehydrogenase). RESULTS: We identified six novel, heterozygous variants in the AMT gene with three predicted to be deleterious to AMT function (p.Val7Leu, p.Pro251Arg, and p.Val380Met). Five extremely rare, known heterozygous variants were found in the AMT gene and one in the GLDC gene. No novel variants in the exons of the other two GCS genes (DLD and GCSH) were identified. CONCLUSION: We identified novel and rare, known variants in two of the four GCS genes that may contribute to the development of MM. Consistent with previous findings, the current study provides additional support that genetic variations in GCS genes contribute to the risk of NTDs. Birth Defects Research (Part A) 106:847-853, 2016. © 2016 Wiley Periodicals, Inc.

Communicating doses of pediatric liquid medicines to parents/caregivers: a comparison of written dosing directions on prescriptions with labels applied by dispensed pharmacy.

OBJECTIVE: To identify and compare volumetric measures used by healthcare providers in communicating dosing instructions for pediatric liquid prescriptions to parents/caregivers. STUDY DESIGN: Dosing instructions were retrospectively reviewed for the 10 most frequently prescribed liquid medications dispensed from 4 community pharmacies for patients aged ≤ 12 years during a 3-month period. Volumetric measures on original prescriptions (ie, milliliters, teaspoons) were compared with those utilized by the pharmacist on the pharmacy label dispensed to the parent/caregiver. RESULTS: Of 649 prescriptions and corresponding pharmacy labels evaluated, 68% of prescriptions and 62% of pharmacy labels communicated dosing in milliliters, 24% of prescriptions and 29% of pharmacy labels communicated dosing in teaspoonfuls, 7% of prescriptions and 0% of pharmacy labels communicated dosing in other measures (ie, milligrams, cubic centimeters, "dose"), and 25% of dispensed pharmacy labels did not reflect units as written in the prescription. CONCLUSION: Volumetric measures utilized by healthcare professionals in dosing instructions for prescription pediatric oral liquid medications are not consistent. Healthcare professionals and parents/caregivers should be educated on safe dosing practices for liquid pediatric medications. Generalizability to the larger pediatric population may vary depending on pharmacy chain, location, and medications evaluated.

Clinical approach to known and emerging tick-borne infections other than Lyme disease.

PURPOSE OF REVIEW: We review latest developments in knowledge of established and emerging tick-borne infections in the United States other than Lyme borreliosis, emphasizing a clinical and geographic approach to diagnosis and management. RECENT FINDINGS: The incidence of tick-borne diseases in the United States has increased. New tick-borne diseases have emerged and will likely continue to be identified. SUMMARY: Clinicians should maintain suspicion for tick-borne diseases in children with acute infectious illnesses, and consider treating such patients presumptively to prevent complications. Knowledge of common tick vectors in the United States and the infections they transmit will allow pediatricians to appropriately assess and manage patients with tick-borne diseases.

Postpartum Use of Shavari Bar® Improves Breast Milk Output: A Double-Blind, Prospective, Randomized, Controlled Clinical Study.

Background and objectives Appropriate nutrition, along with the establishment of lactation, is of paramount importance for the feeding mother and the growing neonate. Asparagus racemosus, a common name for Shatavari, is a well-known herb that has been used as a galactagogue in traditional Indian culture. It is also referenced in Ayurvedic medicine. Despite multiple formulations available, palatability has been a concern always as Shatavari is very bitter. We have devised a palatable and nutritionally rich formulation of Shatavari with no artificial ingredients. To understand the efficacy, we have conducted this double-blind, prospective, randomized, controlled study to evaluate the effect of oral Shatavari formulation (Shavari Bar®) on breast milk output in postpartum women. Methods A prospective, randomized, parallel-group, double-blind, placebo-controlled study was conducted at two centers in women with gestational age 37 weeks or more who intended to breastfeed. Hundred and four women were screened, of which 78 were randomized to receive either bar containing Shatavari and oats (n=39, study) or an identical placebo bar (n=39, control). All 78 women completed the study, 61 delivered by a lower segment Caesarean section (LSCS), and 17 had a full-term normal vaginal delivery. Time to first noticeable breast fullness was measured and expressed milk volume measurements were done 72 hours after delivery or after consumption of four bars, whichever was later using a standardized breast pump. Comparison between the two groups was analyzed using a t-test. Results Demography and baseline data of patients enrolled were similar in the two groups. The mean total milk volume expressed was higher (p=0.008) with Shavari (64.74 ml) compared to placebo (49.69 ml). The time to breast fullness was shorter (p=0.024) with Shavari (30.49 hours) compared to placebo (38.09 hours). No adverse events were noted in either of the study groups. Global assessment of the satisfaction of mothers with lactation, the well-being of the child, taste, and ease of use was better in the treatment arm than in the placebo arm. Conclusion The use of the Shavari bar can be an effective option in postpartum women to establish early lactation and build confidence in breastfeeding along with nonpharmacological intervention.

Case Report: The Medical and Surgical Management of an Infant With Extreme Prematurity and Fetus-In-Fetu.

Fetus-in-fetu (FIF) is a rare congenital anomaly where a parasitic twin is within the body of a host twin. FIF is reported to occur in 1:500,000 live births. Herein, we report the first case of the medical and surgical treatment of a FIF patient who was born with extreme prematurity at 25-weeks gestation. With the multi-disciplinary coordination of neonatology, surgery, and interventional radiology, the patient was able to achieve a window of medical stability 4 weeks after birth. A decision was made at that time to proceed with an intra-abdominal and perineal resection of the FIF. The FIF was successfully resected and the patient was able to recover from the operation, with eventual discharge from the NICU. In conclusion, extreme prematurity and FIF may be amenable to surgical resection and a multi-disciplinary approach is crucial to achieve the desired outcome.

Parolees' physical closeness to health service providers: a study of California parolees.

We studied a sample of parolees and health service providers in the state of California in 2005-2006 to examine the relative physical closeness to health providers (and the potential demand of these providers) of parolees based on their demographic and prior offending characteristics. Although African-American and Latino parolees have more health providers nearby, these providers have considerably more potential demand. The health providers near long-term prisoners and sex offenders have more potential demand. The results suggest inequity in access to services, as minority parolees and those with greater needs may live near more impacted providers. The results also suggest some differences in access based on rural, suburban, or urban location.

HMGB1 is a bone-active cytokine.

High mobility group box 1 (HMGB1) is a chromatin protein that acts as an immunomodulatory cytokine upon active release from myeloid cells. HMGB1 is also an alarmin, an endogenous molecule released by dying cells that acts to initiate tissue repair. We have previously reported that osteoclasts and osteoblasts release HMGB1 and release by the latter is regulated by parathyroid hormone (PTH), an agent of bone remodeling. A recent study suggests that HMGB1 acts as a chemotactic agent to osteoclasts and osteoblasts during endochondral ossification. To explore the potential impact of HMGB1 in the bone microenvironment and its mechanism of release by osseous cells, we characterized the effects of recombinant protein (rHMGB1) on multiple murine bone cell preparations that together exhibit the various cell phenotypes present in bone. We also inquired whether apoptotic bone cells release HMGB1. rHMGB1 enhanced the RANKL/OPG steady state mRNA ratio and dramatically augmented the release of tumor necrosis factor-alpha (TNFalpha) and interleukin-6 (IL6) in osteoblastogenic bone marrow stromal cell (BMSC) cultures but not in the calvarial-derived MC3T3-E1 cells. Interestingly, rHMGB1 promoted GSK-3beta phosphorylation in MC3T3-E1 cells but not in BMSCs. Apoptotic bone cells released HMGB1, including MLO-Y4 osteocyte-like cells. MLO-Y4 release of HMGB1 was coincident with caspase-3 cleavage. Furthermore, the anti-apoptotic action of PTH on MC3T3-E1 cells correlated with the observed decrease in HMGB1 release. Our data suggest that apoptotic bone cells release HMGB1, that within the marrow HMGB1 is a bone resorption signal, and that intramembraneous and endochondral osteoblasts exhibit differential responses to this cytokine.

Burden of Healthcare-Associated Viral Respiratory Infections in Children's Hospitals.

OBJECTIVE: Although healthcare-associated (HA) viral respiratory infections (VRIs) are common in pediatrics, no benchmark for comparison exists. We aimed to determine, compare, and assess determinants of unit-specific HA-VRI incidence rates in 2 children's hospitals. METHODS: This study was a retrospective comparison of prospective cohorts. The Montreal Children's Hospital and the Cohen Children's Medical Center of New York perform prospective surveillance for HA-VRI using standardized definitions that require the presence of symptoms compatible with VRI and virus detection. Cases detected between April 1, 2010, and March 31, 2013, were identified using surveillance databases. Annual incidence rates were calculated, and a generalized estimating equation model was used to assess determinants of HA-VRI rates. RESULTS: The overall HA-VRI rate during the 3-year study period was significantly higher at Montreal Children's Hospital than that at Cohen Children's Medical Center of New York (1.91 vs 0.80 per 1000 patient-days, respectively; P < .0001). Overall, the HA-VRI incidence rate was lowest in the neonatal intensive care unit. Rates in the pediatric intensive care, oncology, and medical/surgical units were similar. The most common etiology of HA-VRI at both institutions was rhinovirus (49% of cases), followed by parainfluenza virus and respiratory syncytial virus. Hospitals with less than 50% single rooms had HA-VRI rates 1.33 (95% confidence interval, 1.29-1.37) times higher than hospitals with more than 50% single rooms for a given unit type. CONCLUSIONS: HA-VRI rates were substantial but different among 2 children's hospitals. Future studies should examine the effect of HA-VRI and evaluate best practices for preventing such infections.

Two promoters control the mouse Nmp4/CIZ transcription factor gene.

Nmp4/CIZ proteins (nuclear matrix protein 4/cas interacting zinc finger protein) contribute to gene regulation in bone, blood, and testis. In osteoblasts, they govern the magnitude of gene response to osteotropic factors like parathyroid hormone (PTH). Nmp4/CIZ is recurrently involved in acute leukemia and it has been implicated in spermatogenesis. However, these conserved proteins, derived from a single gene, are expressed in numerous tissues indicative of a more generalized housekeeping function in addition to their tissue-specific roles. To address how Nmp4/CIZ expression is governed, we characterized the 5' regulatory region of the mouse Nmp4 gene, located on chromosome 6. Two adjacent promoters P(1) [-2521 nucleotide (nt)/-597 nt] and P(2) (-2521 nt/+1 nt) initiate transcription of alternative first exons (U(1) and U(2)). Both promoters lack TATA and CCAAT boxes but contain initiator sites and CpG islands. Northern analysis revealed expression of both U(1) and U(2) in numerous adult tissues consistent with the constitutive and ubiquitous activity of a housekeeping gene. Sequence analysis identified numerous potential transcription factor-binding sites significant to osteogenesis, hematopoeisis, and gonadal development. The promoters are active in both osteoblast-like cells and in the M12 B-lymphocyte cell line. Low doses of PTH attenuated P(1)/P(2) activity in osteoblast-like cells. The Nmp4/CIZ promoters are autoregulated and deletion analysis identified regions that drive P(1) and P(2) basal activities as well as regions that contain positive and negative regulatory elements affecting transcription. The Nmp4/CIZ promoters comprise a genomic regulatory architecture that supports constitutive expression as well as cell- and tissue-specific regulation.

Pattern of bacterial colonization in a new neonatal intensive care unit and its association with infections in infants.

BACKGROUND: There is paucity of information on the pattern of bacterial colonization of a new neonatal intensive care unit. OBJECTIVE: To study the pattern of bacterial colonization on the environmental surfaces in a new neonatal intensive care unit (NICU) and correlate it with infections in the infants. METHODS: Environmental cultures from the faucets and computer keyboards in the NICU were obtained prospectively every 2 weeks for 1 year. Positive blood, cerebrospinal fluid, and respiratory cultures from the infants in the NICU were also obtained. RESULTS: A total of 175 swab cultures was collected, which were sterile for initial 6-week period. Subsequently, 31 cultures grew microbes: 26 (83.8%) from the faucets and 5 (16.2%) from the computers keyboard (P < .001). Of the 48 positive blood cultures in NICU patients, 6 (12.5%) matched the organism growing from the surveillance sites, but the correlation was not significant (P = .076). None of the 31 positive respiratory cultures and 1 positive cerebrospinal fluid culture correlated to the organisms grown from the NICU environment. CONCLUSION: The environment was colonized after an initial period of sterile cultures in a new NICU. Once colonized, they can persist, increasing the risk of developing resistance to antibiotics. They did not correlate with the positive cultures from the infants admitted to the NICU during the study period.

Addressing the workforce crisis: the professional aspirations of pharmacy students in Ghana.

OBJECTIVE: A lack of skilled health professionals, and net migration from developing to more developed countries, are widely recognised as barriers to the delivery of effective health care. However, few studies have investigated this issue from the perspective of pharmacists, although they are increasingly viewed as a potentially valuable and underexploited health care resource. The objectives of this study were to examine the professional aspirations and perceived opportunities of final year pharmacy students in a developing country; and consider what developments may encourage them to remain in, and contribute to, health care in their home country. METHOD: Final year pharmacy students from the Faculty of Pharmacy, KNUST, Kumasi, Ghana, were randomly selected and invited to participate in in-depth interviews. These were audio-recorded (with permission of respondents) and transcribed verbatim to enable a qualitative analysis. MAIN OUTCOME MEASURE: professional aspirations, and perceived opportunities and barriers to their achievement in Ghana and abroad. Results Participants viewed themselves, and wished to be viewed by others, as health professionals. They described a commitment to applying their clinical knowledge and to education beyond their first degree. However, they identified significant barriers to the achievement of professional aspirations in Ghana, which would diminish their opportunities to contribute to health care. Whilst most students expressed the expectation or desire to travel at some point, usually early, in their career, they all demonstrated a commitment to their country and stated a wish to return. CONCLUSION: Overall the study highlighted prospective pharmacists in Ghana as ambitious, committed potential health professionals. The study indicates that a lack of attention by policy makers and professional bodies to ways of exploiting the contribution of pharmacists to public health, may represent a lost potential human resource for health in developing countries.

Nmp4/CIZ contributes to fluid shear stress induced MMP-13 gene induction in osteoblasts.

The expression of matrix metalloproteinase-13 (MMP-13), involved in bone turnover, is elevated in stretched MC3T3-E1 osteoblast-like cells. Strain-mediated forces impact bone remodeling due in large part to the movement of fluid through the canalicular-lacunar network. The resulting fluid shear stress (FSS) over the surface membranes of bone cells initiates bone remodeling. Although the nuclear events mediating putative FSS-induced changes in osteoblast MMP-13 transcription are unknown, previous studies with bone cells suggest an overlap between osteoblast FSS- and PTH-induced signal response pathways. MMP-13 PTH response is regulated by a 110 bp 5' regulatory region, conserved across the mouse, rat, and human genes, that supports the binding of numerous transcription factors including Runx2, c-fos/c-jun, Ets-1, and nuclear matrix protein 4/cas interacting zinc finger protein (Nmp4/CIZ) a nucleocytoplasmic shuttling trans-acting protein that attenuates PTH-driven transcription. Nmp4/CIZ also binds p130(cas), an adaptor protein implicated in mechanotransduction. Here we sought to determine whether Nmp4/CIZ contributes to FSS-induced changes in MMP-13 transcription. FSS (12 dynes/cm(2), 3-5 h) increased MMP-13 promoter-reporter activity approximately two-fold in MC3T3-E1 osteoblast-like cells attended by a comparable increase in mRNA expression. This was accompanied by a decrease in Nmp4/CIZ binding to its cis-element within the PTH response region, the mutation of which abrogated the MMP-13 response to FSS. Interestingly, FSS enhanced Nmp4/CIZ promoter activity and induced p130(cas) nuclear translocation. We conclude that the PTH regulatory region of MMP-13 also contributes to FSS response and that Nmp4/CIZ plays similar but distinct roles in mediating hormone- and FSS-driven induction of MMP-13 in bone cells.

HMGB1 expression and release by bone cells.

Immune and bone cells are functionally coupled by pro-inflammatory cytokine intercellular signaling networks common to both tissues and their crosstalk may contribute to the etiologies of some immune-associated bone pathologies. For example, the receptor activator of NF-kappaB ligand (RANKL)/osteoprotegerin (OPG)/receptor activator of NF-kappaB (RANK) signaling axis plays a critical role in dendritic cell (DC) function as well as bone remodeling. The expression of RANKL by immune cells may contribute to bone loss in periodontitis, arthritis, and multiple myeloma. A recent discovery reveals that DCs release the chromatin protein high mobility group box 1 (HMGB1) as a potent immunomodulatory cytokine mediating the interaction between DCs and T-cells, via HMGB1 binding to the membrane receptor for advanced glycation end products (RAGE). To determine whether osteoblasts or osteoclasts express and/or release HMGB1 into the bone microenvironment, we analyzed tissue, cells, and culture media for the presence of this molecule. Our immunohistochemical and immunocytochemical analyses demonstrate HMGB1 expression in primary osteoblasts and osteoclasts and that both cells express RAGE. HMGB1 is recoverable in the media of primary osteoblast cultures and cultures of isolated osteoclast precursors and osteoclasts. Parathyroid hormone (PTH), a regulator of bone remodeling, attenuates HMGB1 release in cultures of primary osteoblasts and MC3T3-E1 osteoblast-like cells but augments this release in the rat osteosarcoma cell line UMR 106-01, both responses primarily via activation of adenylyl cyclase. PTH-induced HMGB1 discharge by UMR cells exhibits similar release kinetics as reported for activated macrophages. These data confirm the presence of the HMGB1/RAGE signaling axis in bone.

Reversal of diabetes in the rat by injection of hematopoietic stem cells infected with recombinant adeno-associated virus containing the preproinsulin II gene.

AIM: To study the effect of injecting hematopoietic stem cells containing the preproinsulin gene II (rI2) via recombinant adeno-associated virus (rAAV) into normal and streptozotocin-diabetic rats. METHODS: rI2 was transfected into rat hematopoietic stem cells using rAAV vector. Stem cells were injected by intravenous route into normal and STZ-induced diabetic rats to study blood sugar and expression of rI2 in various tissues. The pLP-1 recombinant plasmid containing rI2 (vLP-1) was engineered as previously described. Bone marrow from female Wistar-Furth rats was enriched for stem cells by using plastic adherence and monoclonal antirat CD3 and CD45 RA to deplete T and B cells. The remaining cells were exposed to vLP-1 (multiplicity of infection MOI =50:1 or 100:1) for 2 h. Approximately ten million exposed stem cells were injected by intravenous route into each animal; there were four groups: normal animals at MOI 50:1 (group 1) or MOI 100:1 (group 2); group 3 animals (n = 9) were streptozotocin-induced diabetic animals at MOI 100:1. Animals that showed reversal of diabetes from group 3 were sacrificed for study of gene expression at weeks 1, 2, and 6, respectively. Control diabetic animals did not receive stem cells or virus constituted group 4. Expression of rI2 was analyzed by RT-PCR and Southern analyses. RESULTS: Despite introduction of insulin gene, groups 1 and 2 had blood sugar concentrations that remained within normal levels, while 3 of 9 animals in group 3 showed reversal of diabetes; using RT-PCR,group 1 expressed rI2 in liver, spleen, thymus, brain, and heart at week 1 only. In group 2, rI2 was seen in the thymus up to 6 weeks; in diabetic animals (group 3) rI2 was seen in liver, bone marrow, spleen, thymus, and peripheral blood lymphocytes at week 2 and in thymus and lymphocytes at week 6. CONCLUSIONS: We have shown that (1) rAAV is a useful vector for transferring rI2 into rat hematopoietic stem cells; (2) normal animals remained euglycemic after injection of stem cells containing rI2 despite identification in various tissues suggesting autoregulation, and (3) short-term reversal of diabetes was achieved in some animals by injection of stem cells containing rI2.